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  • Machine learning techniques for protein function prediction
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-11-14
    Rosalin Bonetta; Gianluca Valentino

    Proteins play important roles in living organisms, and their function is directly linked with their structure. Due to the growing gap between the number of proteins being discovered and their functional characterization (in particular as a result of experimental limitations), reliable prediction of protein function through computational means has become crucial. This paper reviews the machine learning techniques used in the literature, following their evolution from simple algorithms such as logistic regression to more advanced methods like support vector machines and modern deep neural networks. Hyperparameter optimization methods adopted to boost prediction performance are presented. In parallel, the metamorphosis in the features used by these algorithms from classical physicochemical properties and amino acid composition, up to text‐derived features from biomedical literature and learned feature representations using autoencoders, together with feature selection and dimensionality reduction techniques, are also reviewed. The success stories in the application of these techniques to both general and specific protein function prediction are discussed.

    更新日期:2020-01-24
  • In silico design and molecular basis for the selectivity of Olinone toward the first over the second bromodomain of BRD4
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-21
    Yoel Rodríguez; Guillermo Gerona‐Navarro; Roman Osman; Ming‐Ming Zhou

    Bromodomains (BrDs), a conserved structural module in chromatin‐associated proteins, are well known for recognizing ε‐N‐acetyl lysine residues on histones. One of the most relevant BrDs is BRD4, a tandem BrD containing protein (BrD1 and BrD2) that plays a critical role in numerous diseases including cancer. Growing evidence shows that the two BrDs of BRD4 have different biological functions; hence selective ligands that can be used to study their functions are of great interest. Here, as a follow‐up of our previous work, we first provide a detailed characterization study of the in silico rational design of Olinone as part of a series of five tetrahydropyrido indole‐based compounds as BRD4 BrD1 inhibitors. Additionally, we investigated the molecular basis for Olinone's selective recognition by BrD1 over BrD2. Molecular dynamics simulations, free energy calculations, and conformational analyses of the apo‐BRD4‐BrD1|2 and BRD4‐BrD1|2/Olinone complexes showed that Olinone's selectivity is facilitated by five key residues: Leu92 in BrD1|385 in BrD2 of ZA loop, Asn140|433, Asp144|His437 and Asp145|Glu438 of BC loop, and Ile146|Val49 of helix C. Furthermore, the difference in hydrogen bonds number and in mobility of the ZA and BC loops of the acetyl‐lysine binding site between BRD4 BrD1/Olinone and BrD2/Olinone complexes also contribute to the difference in Olinone's binding affinity and selectivity toward BrD1 over BrD2. Altogether, our computer‐aided molecular design techniques can effectively guide the development of small‐molecule BRD4 BrD1 inhibitors, explain their selectivity origin, and further open doors to the design of new therapeutically improved derivatives.

    更新日期:2020-01-24
  • Delineation of a new structural motif involving NHN γ‐turn
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-21
    Jesmita Dhar; Raghuvansh Kishore; Pinak Chakrabarti

    Macromolecules are characterized by distinctive arrangement of hydrogen bonds. Different patterns of hydrogen bonds give rise to distinct and stable structural motifs. An analysis of 4114 non‐redundant protein chains reveals the existence of a three‐residue, (i − 1) to (i + 1), structural motif, having two hydrogen‐bonded five‐membered pseudo rings (the first, an NH···OC involving the first residue, and the second being NH∙∙∙N involving the last two residues), separated by a peptide bond. There could be an additional hydrogen bond between the side‐chain at (i‐1) and the main‐chain NH of (i + 1). The average backbone torsion angles of −76(±21)° and – 12(±17)° at i creates a tight turn in the polypeptide chain, akin to a γ‐turn. Indeed, a search of three‐residue fragments with restriction on the terminal Cα···Cα distance and the existence of the two pseudo rings on either side revealed the presence 14 846 cases of a variant, termed NHN γ‐turn, distinct from the NHO γ‐turn (2032 cases) that has traditionally been characterized by the presence of NHO hydrogen bond linking the terminal main‐chain atoms. As in the latter, the newly identified γ‐turns are also of two types—classical and inverse, occurring in the ratio of 1:6. The propensities of residues to occur in these turns and their secondary structural features have been enumerated. An understanding of these turns would be useful for structure prediction and loop modeling, and may serve as models to represent some of the unfolded state or disordered region in proteins.

    更新日期:2020-01-24
  • Lysosomal acid lipase does not have a propeptide and should not be considered being a proprotein
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-24
    Thea B. Strøm; Terje Vinje; Katrine Bjune; Luís T. da Costa; Jon K. Laerdahl; Trond P. Leren

    Lysosomal acid lipase (LAL) plays an important role in lipid metabolism by performing hydrolysis of triglycerides and cholesteryl esters in the lysosome. Based upon characteristics of LAL purified from human liver, it has been proposed that LAL is a proprotein with a 55 residue propeptide that may be essential for proper folding, intracellular transport, or enzymatic function. However, the biological significance of such a propeptide has not been fully elucidated. In this study, we have performed a series of studies in cultured HepG2 and HeLa cells to determine the role of the putative propeptide. However, by Western blot analysis and subcellular fractionation, we have not been able to identify a cleaved LAL lacking the N‐terminal 55 residues. Moreover, mutating residues surrounding the putative cleavage site at Lys76↓ in order to disrupt a proteinase recognition sequence, did not affect LAL activity. Furthermore, forcing cleavage at Lys76↓ by introducing the optimal furin cleavage site RRRR↓EL between residues 76 and 77, did not affect LAL activity. These data, in addition to bioinformatics analyses, indicate that LAL is not a proprotein. Thus, it is possible that the previously reported cleavage at Lys76↓ could have resulted from exposure to proteolytic enzymes during the multistep purification procedure.

    更新日期:2020-01-24
  • Experimental comparison of energy landscape features of ubiquitin family proteins
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-16
    Tathagata Nandi; Anju Yadav; Sri Rama Koti Ainavarapu

    Small ubiquitin‐related modifiers (SUMO1 and SUMO2) are ubiquitin family proteins, structurally similar to ubiquitin, differing in terms of their amino acid sequence and functions. Therefore, they provide a great platform for investigating sequence‐structure‐stability‐function relationship. Here, we used chemical denaturation in comparing the folding‐unfolding pathways of the SUMO proteins with their structural homologue ubiquitin (UF45W‐pseudo wild‐type [WT] tryptophan variant) with structurally analogous tryptophan mutations (SUMO1 [S1F66W], SUMO2 [S2F62W]). Equilibrium denaturation studies report that ubiquitin is the most stable protein among the three. The observed denaturant‐dependent folding rates of SUMOs are much lower than ubiquitin and primarily exhibit a two‐state folding pathway unlike ubiquitin, which has a kinetic folding intermediate. We hypothesize that, as SUMO proteins start off as slow folders, they avoid stabilizing their folding intermediates and the presence of which might further slow‐down their folding rates. The denaturant‐dependent unfolding of ubiquitin is the fastest, followed by SUMO2, and slowest for SUMO1. However, the spontaneous unfolding rate constant is the lowest for ubiquitin (~40 times), and similar for SUMOs. This correlation between thermodynamic stability and kinetic stability is achieved by having different unfolding transition state positions with respect to the solvent‐accessible surface area, as quantified by the Tanford β u values: ubiquitin (0.42) > SUMO2 (0.20) > SUMO1 (0.16). The results presented here highlight the unique energy landscape features which help in optimizing the folding‐unfolding rates within a structurally homologous protein family.

    更新日期:2020-01-24
  • Function2Form Bridge—Toward synthetic protein holistic performance prediction
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-29
    Venkata V. B. Yallapragada; Sidney P. Walker; Ciaran Devoy; Stephen Buckley; Yensi Flores; Mark Tangney

    Protein engineering and synthetic biology stand to benefit immensely from recent advances in silico tools for structural and functional analyses of proteins. In the context of designing novel proteins, current in silico tools inform the user on individual parameters of a query protein, with output scores/metrics unique to each parameter. In reality, proteins feature multiple “parts”/functions and modification of a protein aimed at altering a given part, typically has collateral impact on other protein parts. A system for prediction of the combined effect of design parameters on the overall performance of the final protein does not exist. Function2Form Bridge (F2F‐Bridge) attempts to address this by combining the scores of different design parameters pertaining to the protein being analyzed into a single easily interpreted output describing overall performance. The strategy comprises of (a) a mathematical strategy combining data from a myriad of in silico tools into an OP‐score (a singular score informing on a user‐defined overall performance) and (b) the F2F Plot, a graphical means of informing the wetlab biologist holistically on designed construct suitability in the context of multiple parameters, highlighting scope for improvement. F2F predictive output was compared with wetlab data from a range of synthetic proteins designed, built, and tested for this study. Statistical/machine learning approaches for predicting overall performance, for use alongside the F2F plot, were also examined. Comparisons between wetlab performance and F2F predictions demonstrated close and reliable correlations. This user‐friendly strategy represents a pivotal enabler in increasing the accessibility of synthetic protein building and de novo protein design.

    更新日期:2020-01-24
  • Accessory mutations balance the marginal stability of the HIV‐1 protease in drug resistance
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-21
    Thomas R. Weikl; Bahram Hemmateenejad

    The HIV‐1 protease is a major target of inhibitor drugs in AIDS therapies. The therapies are impaired by mutations of the HIV‐1 protease that can lead to resistance to protease inhibitors. These mutations are classified into major mutations, which usually occur first and clearly reduce the susceptibility to protease inhibitors, and minor, accessory mutations that occur later and individually do not substantially affect the susceptibility to inhibitors. Major mutations are predominantly located in the active site of the HIV‐1 protease and can directly interfere with inhibitor binding. Minor mutations, in contrast, are typically located distal to the active site. A central question is how these distal mutations contribute to resistance development. In this article, we present a systematic computational investigation of stability changes caused by major and minor mutations of the HIV‐1 protease. As most small single‐domain proteins, the HIV‐1 protease is only marginally stable. Mutations that destabilize the folded, active state of the protease therefore can shift the conformational equilibrium towards the unfolded, inactive state. We find that the most frequent major mutations destabilize the HIV‐1 protease, whereas roughly half of the frequent minor mutations are stabilizing. An analysis of protease sequences from patients in treatment indicates that the stabilizing minor mutations are frequently correlated with destabilizing major mutations, and that highly resistant HIV‐1 proteases exhibit significant fractions of stabilizing mutations. Our results thus indicate a central role of minor mutations in balancing the marginal stability of the protease against the destabilization induced by the most frequent major mutations.

    更新日期:2020-01-24
  • The unusual conformation of cross‐strand disulfide bonds is critical to the stability of β‐hairpin peptides
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-16
    Evelyne Deplazes; Yanni K.‐Y. Chin; Glenn F. King; Ricardo L. Mancera

    The cross‐strand disulfides (CSDs) found in β‐hairpin antimicrobial peptides (β‐AMPs) show a unique disulfide geometry that is characterized by unusual torsion angles and a short Cα‐Cα distance. While the sequence and disulfide bond connectivity of disulfide‐rich peptides is well studied, much less is known about the disulfide geometry found in CSDs and their role in the stability of β‐AMPs. To address this, we solved the nuclear magnetic resonance (NMR) structure of the β‐AMP gomesin (Gm) at 278, 298, and 310 K, examined the disulfide bond geometry of over 800 disulfide‐rich peptides, and carried out extensive molecular dynamics (MD) simulation of the peptides Gm and protegrin. The NMR data suggests Cα‐Cα distances characteristic for CSDs are independent of temperature. Analysis of disulfide‐rich peptides from the Protein Data Bank revealed that right‐handed and left‐handed rotamers are equally likely in CSDs. The previously reported preference for right‐handed rotamers was likely biased by restricting the analysis to peptides and proteins solved using X‐ray crystallography. Furthermore, data from MD simulations showed that the short Cα‐Cα distance is critical for the stability of these peptides. The unique disulfide geometry of CSDs poses a challenge to biomolecular force fields and to retain the stability of β‐hairpin fold over long simulation times, restraints on the torsion angles might be required.

    更新日期:2020-01-24
  • Geometrical characterization of T cell receptor binding modes reveals class‐specific binding to maximize access to antigen
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-21
    Nishant K. Singh; Esam T. Abualrous; Cory M. Ayres; Frank Noé; Ragul Gowthaman; Brian G. Pierce; Brian M. Baker

    Recognition of antigenic peptides bound to major histocompatibility complex (MHC) proteins by αβ T cell receptors (TCRs) is a hallmark of T cell mediated immunity. Recent data suggest that variations in TCR binding geometry may influence T cell signaling, which could help explain outliers in relationships between physical parameters such as TCR‐pMHC binding affinity and T cell function. Traditionally, TCR binding geometry has been described with simple descriptors such as the crossing angle, which quantifies what has become known as the TCR's diagonal binding mode. However, these descriptors often fail to reveal distinctions in binding geometry that are apparent through visual inspection. To provide a better framework for relating TCR structure to T cell function, we developed a comprehensive system for quantifying the geometries of how TCRs bind peptide/MHC complexes. We show that our system can discern differences not clearly revealed by more common methods. As an example of its potential to impact biology, we used it to reveal differences in how TCRs bind class I and class II peptide/MHC complexes, which we show allow the TCR to maximize access to and “read out” the peptide antigen. We anticipate our system will be of use in not only exploring these and other details of TCR‐peptide/MHC binding interactions, but also addressing questions about how TCR binding geometry relates to T cell function, as well as modeling structural properties of class I and class II TCR‐peptide/MHC complexes from sequence information. The system is available at https://tcr3d.ibbr.umd.edu/tcr_com or for download as a script.

    更新日期:2020-01-24
  • Molecular basis of drug resistance in smoothened receptor: An in silico study of protein resistivity and specificity
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-16
    Noopur Sinha; Saikat Chowdhury; Ram Rup Sarkar

    Smoothened (SMO) antagonist Vismodegib effectively inhibits the Hedgehog pathway in proliferating cancer cells. In early stage of treatment, Vismodegib exhibited promising outcomes to regress the tumors cells, but ultimately relapsed due to the drug resistive mutations in SMO mostly occurring before (primary mutations G497W) or after (acquired mutations D473H/Y) anti‐SMO therapy. This study investigates the unprecedented insights of structural and functional mechanism hindering the binding of Vismodegib with sensitive and resistant mutant variants of SMO (SMOMut). Along with the basic dynamic understanding of Vismodegib‐SMO complexes, network propagation theory based on heat diffusion principles is first time applied here to identify the modules of residues influenced by the individual mutations. The allosteric modulation by GLY497 residue in Vismodegib bound SMO wild‐type (SMOWT) conformation depicts the interconnections of intermediate residues of SMO with the atom of Vismodegib and identify two important motifs (E‐X‐P‐L) and (Q‐A‐N‐V‐T‐I‐G) mediating this allosteric regulation. In this study a novel computational framework based on the heat diffusion principle is also developed, which identify significant residues of allosteric site causing drug resistivity in SMOMut. This framework could also be useful for assessing the potential allosteric sites of different other proteins. Moreover, previously reported novel inhibitor “ZINC12368305,” which is proven to make an energetically favorable complex with SMOWT is chosen as a control sample to assess the impact of receptor mutation on its binding and subsequently identify the important factors that govern binding disparity between Vismodegib and ZINC12368305 bound SMOWT/Mut conformations.

    更新日期:2020-01-24
  • Protonation state of the selectivity filter of bacterial voltage‐gated sodium channels is modulated by ions
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-11-12
    Ana Damjanovic; Ada Y. Chen; Robert L. Rosenberg; Daniel R. Roe; Xiongwu Wu; Bernard R. Brooks

    The selectivity filter (SF) of bacterial voltage‐gated sodium channels consists of four glutamate residues arranged in a C4 symmetry. The protonation state population of this tetrad is unclear. To address this question, we simulate the pore domain of bacterial voltage‐gated sodium channel of Magnetococcus sp. (NavMs) through constant pH methodology in explicit solvent and free energy perturbation calculations. We find that at physiological pH the fully deprotonated as well as singly and doubly protonated states of the SF appear feasible, and that the calculated pKa decreases with each additional bound ion, suggesting that a decrease in the number of ions in the pore can lead to protonation of the SF. Previous molecular dynamics simulations have suggested that protonation can lead to a decrease in the conductance, but no pKa calculations were performed. We confirm a decreased ionic population of the pore with protonation, and also observe structural symmetry breaking triggered by protonation; the SF of the deprotonated channel is closest to the C4 symmetry observed in crystal structures of the open state, while the SF of protonated states display greater levels of asymmetry which could lead to transition to the inactivated state which possesses a C2 symmetry in the crystal structure. We speculate that the decrease in the number of ions near the mouth of the channel, due to either random fluctuations or ion depletion due to conduction, could be a self‐regulatory mechanism resulting in a nonconducting state that functionally resembles inactivated states.

    更新日期:2020-01-24
  • VH‐VL interdomain dynamics observed by computer simulations and NMR
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-14
    Monica L. Fernández‐Quintero; Valentin J. Hoerschinger; Leonida M. Lamp; Alexander Bujotzek; Guy Georges; Klaus R. Liedl

    The relative orientation of the two variable domains, VH and VL, influences the shape of the antigen binding site, that is, the paratope, and is essential to understand antigen specificity. ABangle characterizes the VH‐VL orientation by using five angles and a distance and compares it to other known structures. Molecular dynamics simulations of antibody variable domains (Fvs) reveal fluctuations in the relative domain orientations. The observed dynamics between these domains are confirmed by NMR experiments on a single‐chain variable fragment antibody (scFv) in complex with IL‐1β and an antigen‐binding fragment (Fab). The variability of these relative domain orientations can be interpreted as a structural feature of antibodies, which increases the antibody repertoire significantly and can enlarge the number of possible binding partners substantially. The movements of the VH and VL domains are well sampled with molecular dynamics simulations and are in agreement with the NMR ensemble. Fast Fourier transformation of the ABangle metrics allows to assign timescales of 0.1‐10 GHz to the fastest collective interdomain movements. The results clearly show the necessity of dynamics to understand and characterize the favorable orientations of the VH and VL domains implying a considerable binding interface flexibility and reveal in all antibody fragments (Fab, scFv, and Fv) very similar VH‐VL interdomain variations comparable to the distributions observed for known X‐ray structures of antibodies.

    更新日期:2020-01-15
  • Modeling protein‐protein, protein‐peptide, and protein‐oligosaccharide complexes: CAPRI 7th edition
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-10
    Marc F. Lensink; Nurul Nadzirin; Sameer Velankar; Shoshana J. Wodak

    We present the seventh report on the performance of methods for predicting the atomic resolution structures of protein complexes offered as targets to the community‐wide initiative on the Critical Assessment of Predicted Interactions. Performance was evaluated on the basis of 36 114 models of protein complexes submitted by 57 groups—including 13 automatic servers—in prediction rounds held during the years 2016 to 2019 for eight protein‐protein, three protein‐peptide, and five protein‐oligosaccharide targets with different length ligands. Six of the protein‐protein targets represented challenging hetero‐complexes, due to factors such as availability of distantly related templates for the individual subunits, or for the full complex, inter‐domain flexibility, conformational adjustments at the binding region, or the multi‐component nature of the complex. The main challenge for the protein‐peptide and protein‐oligosaccharide complexes was to accurately model the ligand conformation and its interactions at the interface. Encouragingly, models of acceptable quality, or better, were obtained for a total of six protein‐protein complexes, which included four of the challenging hetero‐complexes and a homo‐decamer. But fewer of these targets were predicted with medium or higher accuracy. High accuracy models were obtained for two of the three protein‐peptide targets, and for one of the protein‐oligosaccharide targets. The remaining protein‐sugar targets were predicted with medium accuracy. Our analysis indicates that progress in predicting increasingly challenging and diverse types of targets is due to closer integration of template‐based modeling techniques with docking, scoring, and model refinement procedures, and to significant incremental improvements in the underlying methodologies.

    更新日期:2020-01-10
  • An overview of data‐driven HADDOCK strategies in CAPRI rounds 38‐45
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-07
    Panagiotis I. Koukos; Jorge Roel‐Touris; Francesco Ambrosetti; Cunliang Geng; Jörg Schaarschmidt; Mikael E. Trellet; Adrien S. J. Melquiond; Li C. Xue; Rodrigo V. Honorato; Irina Moreira; Zeynep Kurkcuoglu; Anna Vangone; Alexandre M. J. J. Bonvin

    Our information‐driven docking approach HADDOCK has demonstrated a sustained performance since the start of its participation to CAPRI. This is due, in part, to its ability to integrate data into the modeling process, and to the robustness of its scoring function. We participated in CAPRI both as server and manual predictors. In CAPRI rounds 38‐45, we have used various strategies depending on the available information. These ranged from imposing restraints to a few residues identified from literature as being important for the interaction, to binding pockets identified from homologous complexes or template‐based refinement/CA‐CA restraint‐guided docking from identified templates. When relevant, symmetry restraints were used to limit the conformational sampling. We also tested for a large decamer target a new implementation of the MARTINI coarse‐grained force field in HADDOCK. Overall, we obtained acceptable or better predictions for 13 and 11 server and manual submissions, respectively, out of the 22 interfaces. Our server performance (acceptable or higher‐quality models when considering the top 10) was better (59%) than the manual (50%) one, in which we typically experiment with various combinations of protocols and data sources. Again, our simple scoring function based on a linear combination of intermolecular van der Waals and electrostatic energies and an empirical desolvation term demonstrated a good performance in the scoring experiment with a 63% success rate across all 22 interfaces. An analysis of model quality indicates that, while we are consistently performing well in generating acceptable models, there is room for improvement for generating/identifying higher quality models.

    更新日期:2020-01-07
  • Modeling beta‐sheet peptide‐protein interactions: Rosetta FlexPepDock in CAPRI rounds 38‐45
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-06
    Alisa Khramushin; Orly Marcu; Nawsad Alam; Orly Shimony; Dzmitry Padhorny; Emiliano Brini; Ken A. Dill; Sandor Vajda; Dima Kozakov; Ora Schueler‐Furman

    Peptide‐protein docking is challenging due to the considerable conformational freedom of the peptide. CAPRI rounds 38‐45 included two peptide‐protein interactions, both characterized by a peptide forming an additional beta strand of a beta sheet in the receptor. Using the Rosetta FlexPepDock peptide docking protocol we generated top‐performing, high‐accuracy models for targets 134 and 135, involving an interaction between a peptide derived from L‐MAG with DLC8. In addition, we were able to generate the only medium‐accuracy models for a particularly challenging target, T121. In contrast to the classical peptide‐mediated interaction, in which receptor side chains contact both peptide backbone and side chains, beta‐sheet complementation involves a major contribution to binding by hydrogen bonds between main chain atoms. To establish how binding affinity and specificity are established in this special class of peptide‐protein interactions, we extracted PeptiDBeta, a benchmark of solved structures of different protein domains that are bound by peptides via beta‐sheet complementation, and tested our protocol for global peptide‐docking PIPER‐FlexPepDock on this dataset. We find that the beta‐strand part of the peptide is sufficient to generate approximate and even high resolution models of many interactions, but inclusion of adjacent motif residues often provides additional information necessary to achieve high resolution model quality.

    更新日期:2020-01-06
  • ProDCoNN: Protein design using a convolutional neural network
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-06
    Yuan Zhang; Yang Chen; Chenran Wang; Chun‐Chao Lo; Xiuwen Liu; Wei Wu; Jinfeng Zhang

    Designing protein sequences that fold to a given three‐dimensional (3D) structure has long been a challenging problem in computational structural biology with significant theoretical and practical implications. In this study, we first formulated this problem as predicting the residue type given the 3D structural environment around the C α atom of a residue, which is repeated for each residue of a protein. We designed a nine‐layer 3D deep convolutional neural network (CNN) that takes as input a gridded box with the atomic coordinates and types around a residue. Several CNN layers were designed to capture structure information at different scales, such as bond lengths, bond angles, torsion angles, and secondary structures. Trained on a very large number of protein structures, the method, called ProDCoNN (protein design with CNN), achieved state‐of‐the‐art performance when tested on large numbers of test proteins and benchmark datasets.

    更新日期:2020-01-06
  • The impact of different mutations at arginine141 on the structure, subunit exchange dynamics and chaperone activity of Hsp16.3
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2020-01-06
    Alok Kumar Panda; Ayon Chakraborty; Sandip Kumar Nandi; Ashis Biswas

    Hsp16.3, a molecular chaperone, plays a vital role in the growth and survival of Mycobacterium tuberculosis inside the host. We previously reported that deletion of three amino acid residues (142STN144) from C‐terminal extension (CTE) of Hsp16.3 triggers its structural perturbation and increases its chaperone activity, which reaches its apex upon the deletion of its entire CTE (141RSTN144). Thus, we hypothesized that Arg141 (R141) and Ser142 (S142) in the CTE of Hsp16.3 possibly hold the key in maintaining its native‐like structure and chaperone activity. To test this hypothesis, we generated two deletion mutants in which R141 and S142 were deleted individually (Hsp16.3ΔR141 and Hsp16.3ΔS142) and three substitution mutants in which R141 was replaced by lysine (Hsp16.3R141K), alanine (Hsp16.3R141A), and glutamic acid (Hsp16.3R141E), respectively. Hsp16.3ΔS142 or Hsp16.3R141K mutant has native‐like structure and chaperone activity. Deletion of R141 from the CTE (Hsp16.3ΔR141) perturbs the secondary and tertiary structure, lowers the subunit exchange dynamics and decreases the chaperone activity of Hsp16.3. But, the substitution of R141 with alanine (Hsp16.3R141A) or glutamic acid (Hsp16.3R141E) perturbs its secondary and tertiary structure. Surprisingly, such charge tampering of R141 enhances the subunit exchange dynamics and chaperone activity of Hsp16.3. Interestingly, neither the deletion of R141/S142 nor the substitution of R141 with lysine, alanine and glutamic acid affects the oligomeric mass/size of Hsp16.3. Overall, our study suggests that R141 (especially the positive charge on R141) plays a crucial role in maintaining the native‐like structure as well as in regulating subunit exchange dynamics and chaperone activity of Hsp16.3.

    更新日期:2020-01-06
  • Characterization of perdeuterated high‐potential iron‐sulfur protein with high‐resolution X‐ray crystallography
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-10
    Yuya Hanazono; Kazuki Takeda; Kunio Miki

    Perdeuteration in neutron crystallography is an effective method for determining the positions of hydrogen atoms in proteins. However, there is shortage of evidence that the high‐resolution details of perdeuterated proteins are consistent with those of the nondeuterated proteins. In this study, we determined the X‐ray structure of perdeuterated high‐potential iron‐sulfur protein (HiPIP) at a high resolution of 0.85 å resolution. The comparison of the nondeuterated and perdeuterated structures of HiPIP revealed slight differences between the two structures. The spectroscopic and spectroelectrochemical studies also showed that perdeuterated HiPIP has approximately the same characteristics as nondeuterated HiPIP. These results further emphasize the suitability of using perdeuterated proteins in the high‐resolution neutron crystallography.

    更新日期:2020-01-04
  • Exploring alternative catalytic mechanisms of the Cas9 HNH domain
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-09-06
    Li Na Zhao; Dibyendu Mondal; Arieh Warshel

    Understanding the reaction mechanism of CRISPR‐associated protein 9 (Cas9) is crucial for the application of programmable gene editing. Despite the availability of the structures of Cas9 in apo‐ and substrate‐bound forms, the catalytically active structure is still unclear. Our first attempt to explore the catalytic mechanism of Cas9 HNH domain has been based on the reasonable assumption that we are dealing with the same mechanism as endonuclease VII, including the assumption that the catalytic water is in the first shell of the Mg2+. Trying this mechanism with the cryo‐EM structure forced us to induce significant structural change driven by the movement of K848 (or other positively charged residue) close to the active site to facilitate the proton transfer step. In the present study, we explore a second reaction mechanism where the catalytic water is in the second shell of the Mg2+ and assume that the cryo‐EM structure by itself is a suitable representation of a catalytic‐ready structure. The alternative mechanism indicates that if the active water is from the second shell, then the calculated reaction barrier is lower compared with the corresponding barrier when the water comes from the first shell.

    更新日期:2020-01-04
  • Comparative dynamics of tropomyosin in vertebrates and invertebrates
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-28
    Jose K. James; Vikas Nanda

    Tropomyosin (Tpm) is an extended α‐helical coiled‐coil homodimer that regulates actinomyosin interactions in muscle. Molecular simulations of four Tpms, two from the vertebrate class Mammalia (rat and pig), and two from the invertebrate class Malacostraca (shrimp and lobster), showed that despite extensive sequence and structural homology across metazoans, dynamic behavior—particularly long‐range structural fluctuations—were clearly distinct. Vertebrate Tpms were more flexible and sampled complex, multi‐state conformational landscapes. Invertebrate Tpms were more rigid, sampling a highly constrained harmonic landscape. Filtering of trajectories by principle component analysis into essential subspaces showed significant overlap within but not between phyla. In vertebrate Tpms, hinge‐regions decoupled long‐range interhelical motions and suggested distinct domains. In contrast, crustacean Tpms did not exhibit long‐range dynamic correlations—behaving more like a single rigid rod on the nanosecond time scale. These observations suggest there may be divergent mechanisms for Tpm binding to actin filaments, where conformational flexibility in mammalian Tpm allows a preorganized shape complementary to the filament surface, and where rigidity in the crustacean Tpm requires concerted bending and binding.

    更新日期:2020-01-04
  • Phylogenetic spread of sequence data affects fitness of consensus enzymes: Insights from triosephosphate isomerase
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-09-13
    Venuka Durani Goyal; Brandon J. Sullivan; Thomas J. Magliery

    The concept of consensus in multiple sequence alignments (MSAs) has been used to design and engineer proteins previously with some success. However, consensus design implicitly assumes that all amino acid positions function independently, whereas in reality, the amino acids in a protein interact with each other and work cooperatively to produce the optimum structure required for its function. Correlation analysis is a tool that can capture the effect of such interactions. In a previously published study, we made consensus variants of the triosephosphate isomerase (TIM) protein using MSAs that included sequences form both prokaryotic and eukaryotic organisms. These variants were not completely native‐like and were also surprisingly different from each other in terms of oligomeric state, structural dynamics, and activity. Extensive correlation analysis of the TIM database has revealed some clues about factors leading to the unusual behavior of the previously constructed consensus proteins. Among other things, we have found that the more ill‐behaved consensus mutant had more broken correlations than the better‐behaved consensus variant. Moreover, we report three correlation and phylogeny‐based consensus variants of TIM. These variants were more native‐like than the previous consensus mutants and considerably more stable than a wild‐type TIM from a mesophilic organism. This study highlights the importance of choosing the appropriate diversity of MSA for consensus analysis and provides information that can be used to engineer stable enzymes.

    更新日期:2020-01-04
  • Boosting phosphorylation site prediction with sequence feature‐based machine learning
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-22
    Shyantani Maiti; Atif Hassan; Pralay Mitra

    Protein phosphorylation is one of the essential posttranslation modifications playing a vital role in the regulation of many fundamental cellular processes. We propose a LightGBM‐based computational approach that uses evolutionary, geometric, sequence environment, and amino acid‐specific features to decipher phosphate binding sites from a protein sequence. Our method, while compared with other existing methods on 2429 protein sequences taken from standard Phospho.ELM (P.ELM) benchmark data set featuring 11 organisms reports a higher F1 score = 0.504 (harmonic mean of the precision and recall) and ROC AUC = 0.836 (area under the curve of the receiver operating characteristics). The computation time of our proposed approach is much less than that of the recently developed deep learning‐based framework. Structural analysis on selected protein sequences informs that our prediction is the superset of the phosphorylation sites, as mentioned in P.ELM data set. The foundation of our scheme is manual feature engineering and a decision tree‐based classification. Hence, it is intuitive, and one can interpret the final tree as a set of rules resulting in a deeper understanding of the relationships between biophysical features and phosphorylation sites. Our innovative problem transformation method permits more control over precision and recall as is demonstrated by the fact that if we incorporate output probability of the existing deep learning framework as an additional feature, then our prediction improves (F1 score = 0.546; ROC AUC = 0.849). The implementation of our method can be accessed at http://cse.iitkgp.ac.in/~pralay/resources/PPSBoost/ and is mirrored at https://cosmos.iitkgp.ac.in/PPSBoost.

    更新日期:2020-01-04
  • Pre‐ and post‐docking sampling of conformational changes using ClustENM and HADDOCK for protein‐protein and protein‐DNA systems
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-09-03
    Zeynep Kurkcuoglu; Alexandre M. J. J. Bonvin

    Incorporating the dynamic nature of biomolecules in the modeling of their complexes is a challenge, especially when the extent and direction of the conformational changes taking place upon binding is unknown. Estimating whether the binding of a biomolecule to its partner(s) occurs in a conformational state accessible to its unbound form (“conformational selection”) and/or the binding process induces conformational changes (“induced‐fit”) is another challenge. We propose here a method combining conformational sampling using ClustENM—an elastic network‐based modeling procedure—with docking using HADDOCK, in a framework that incorporates conformational selection and induced‐fit effects upon binding. The extent of the applied deformation is estimated from its energetical costs, inspired from mechanical tensile testing on materials. We applied our pre‐ and post‐docking sampling of conformational changes to the flexible multidomain protein‐protein docking benchmark and a subset of the protein‐DNA docking benchmark. Our ClustENM‐HADDOCK approach produced acceptable to medium quality models in 7/11 and 5/6 cases for the protein‐protein and protein‐DNA complexes, respectively. The conformational selection (sampling prior to docking) has the highest impact on the quality of the docked models for the protein‐protein complexes. The induced‐fit stage of the pipeline (post‐sampling), however, improved the quality of the final models for the protein‐DNA complexes. Compared to previously described strategies to handle conformational changes, ClustENM‐HADDOCK performs better than two‐body docking in protein‐protein cases but worse than a flexible multidomain docking approach. However, it does show a better or similar performance compared to previous protein‐DNA docking approaches, which makes it a suitable alternative.

    更新日期:2020-01-04
  • The characterization of pc‐polylines representing protein backbones
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-04
    Lincong Wang; Yao Zhang; Shuxue Zou

    The backbone of a protein is typically represented as either a C α‐polyline, a three‐dimensional (3D) polyline that passes through the C α atoms, or a tuple of ϕ,ψ pairs while its fold is usually assigned using the 3D topological arrangement of the secondary structure elements (SSEs). It is tricky to obtain the SSE composition for a protein from the C α‐polyline representation while its 3D SSE arrangement is not apparent in the two‐dimensional (2D) ϕ,ψ representation. In this article, we first represent the backbone of a protein as a pc‐polyline that passes through the centers of its peptide planes. We then analyze the pc‐polylines for six different sets of proteins with high quality crystal structures. The results show that SSE composition becomes recognizable in pc‐polyline presentation and consequently the geometrical property of the pc‐polyline of a protein could be used to assign its secondary structure. Furthermore, our analysis finds that for each of the six sets the total length of a pc‐polyline increases linearly with the number of the peptide planes. Interestingly a comparison of the six regression lines shows that they have almost identical slopes but different intercepts. Most interestingly there exist decent linear correlations between the intercepts of the six lines and either the average helix contents or the average sheet contents and between the intercepts and the average backbone hydrogen bonding energetics. Finally, we discuss the implications of the identified correlations for structure classification and protein folding, and the potential applications of pc‐polyline representation to structure prediction and protein design.

    更新日期:2020-01-04
  • The structural details of the interaction of single‐stranded DNA binding protein hSSB2 (NABP1/OBFC2A) with UV‐damaged DNA
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-30
    Teegan Lawson; Serene El‐Kamand; Didier Boucher; Duc Cong Duong; Ruvini Kariawasam; Alexandre M. J. J. Bonvin; Derek J. Richard; Roland Gamsjaeger; Liza Cubeddu

    Single‐stranded DNA‐binding proteins (SSBs) are required for all known DNA metabolic events such as DNA replication, recombination and repair. While a wealth of structural and functional data is available on the essential human SSB, hSSB1 (NABP2/OBFC2B), the close homolog hSSB2 (NABP1/OBFC2A) remains relatively uncharacterized. Both SSBs possess a well‐structured OB (oligonucleotide/oligosaccharide‐binding) domain that is able to recognize single‐stranded DNA (ssDNA) followed by a flexible carboxyl‐tail implicated in the interaction with other proteins. Despite the high sequence similarity of the OB domain, several recent studies have revealed distinct functional differences between hSSB1 and hSSB2. In this study, we show that hSSB2 is able to recognize cyclobutane pyrimidine dimers (CPD) that form in cellular DNA as a consequence of UV damage. Using a combination of biolayer interferometry and NMR, we determine the molecular details of the binding of the OB domain of hSSB2 to CPD‐containing ssDNA, confirming the role of four key aromatic residues in hSSB2 (W59, Y78, W82, and Y89) that are also conserved in hSSB1. Our structural data thus demonstrate that ssDNA recognition by the OB fold of hSSB2 is highly similar to hSSB1, indicating that one SSB may be able to replace the other in any initial ssDNA binding event. However, any subsequent recruitment of other repair proteins most likely depends on the divergent carboxyl‐tail and as such is likely to be different between hSSB1 and hSSB2.

    更新日期:2020-01-04
  • Effect of hydrophobic and hydrogen bonding interactions on the potency of ß‐alanine analogs of G‐protein coupled glucagon receptor inhibitors
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-09-10
    Pushyaraga P. Venugopal; Bratin K. Das; E. Soorya; Debashree Chakraborty

    G‐protein coupled glucagon receptors (GCGRs) play an important role in glucose homeostasis and pathophysiology of Type‐II Diabetes Mellitus (T2DM). The allosteric pocket located at the trans‐membrane domain of GCGR consists of hydrophobic (TM5) and hydrophilic (TM7) units. Hydrophobic interactions with the amino acid residues present at TM5, found to facilitate the favorable orientation of antagonist at GCGR allosteric pocket. A statistically robust and highly predictive 3D‐QSAR model was developed using 58 β‐alanine based GCGR antagonists with significant variation in structure and potency profile. The correlation coefficient (R2) and cross‐validation coefficient (Q2) of the developed model were found to be 0.9981 and 0.8253, respectively at the PLS factor of 8. The analysis of the favorable and unfavorable contribution of different structural features on the glucagon receptor antagonists was done by 3D‐QSAR contour plots. Hydrophobic and hydrogen bonding interactions are found to be main dominating non‐bonding interactions in docking studies. Presence of highest occupied molecular orbital (HOMO) in the polar part and lowest unoccupied molecular orbital (LUMO) in the hydrophobic part of antagonists leads to favorable protein‐ligand interactions. Molecular mechanics/generalized born surface area (MM/GBSA) calculations showed that van der Waals and nonpolar solvation energy terms are crucial components for thermodynamically stable binding of the inhibitors. The binding free energy of highly potent compound was found to be −63.475 kcal/mol; whereas the least active compound exhibited binding energy of −41.097 kcal/mol. Further, five 100 ns molecular dynamics simulation (MD) simulations were done to confirm the stability of the inhibitor‐receptor complex. Outcomes of the present study can serve as the basis for designing improved GCGR antagonists.

    更新日期:2020-01-04
  • MD simulations reveal alternate conformations of the oxyanion hole in the Zika virus NS2B/NS3 protease
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-09-09
    Jinhong Ren; Hyun Lee; Alpa Kotak; Michael E. Johnson

    Recent crystallography studies have shown that the binding site oxyanion hole plays an important role in inhibitor binding, but can exist in two conformations (active/inactive). We have undertaken molecular dynamics (MD) calculations to better understand oxyanion hole dynamics and thermodynamics. We find that the Zika virus (ZIKV) NS2B/NS3 protease maintains a stable closed conformation over multiple 100‐ns conventional MD simulations in both the presence and absence of inhibitors. The S1, S2, and S3 pockets are stable as well. However, in two of eight simulations, the A132‐G133 peptide bond in the binding pocket of S1' spontaneously flips to form a 310‐helix that corresponds to the inactive conformation of the oxyanion hole, and then maintains this conformation until the end of the 100‐ns conventional MD simulations without inversion of the flip. This conformational change affects the S1' pocket in ZIKV NS2B/NS3 protease active site, which is important for small molecule binding. The simulation results provide evidence at the atomic level that the inactive conformation of the oxyanion hole is more favored energetically when no specific interactions are formed between substrate/inhibitor and oxyanion hole residues. Interestingly, however, transition between the active and inactive conformation of the oxyanion hole can be observed by boosting the valley potential in accelerated MD simulations. This supports a proposed induced‐fit mechanism of ZIKV NS2B/NS3 protease from computational methods and provides useful direction to enhance inhibitor binding predictions in structure‐based drug design.

    更新日期:2020-01-04
  • On predicting foldability of a protein from its sequence
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-03
    Mihaly Mezei

    Several properties of amino acid sequences corresponding to proteins that are known to fold are compared to those of randomly generated sequences and to sequences of intrinsically disordered proteins in order to find properties that distinguish folding sequences from the rest. The properties studied included helix and sheet propensities from secondary structure prediction, adjacency correlations, directionality correlations, as well as propensities of all possible triplets and quadruplets. Small differences between known folded and random sequences were observed for the adjacency and directional correlations, and significant differences were seen on the triplet and especially on the quadruplet propensities. Based on the differences in the adjacency, triplet or quadruplet propensities folding scores were defined and used to test the accuracy of foldability prediction based on these statistics. The best predictions were obtained from the quadruplet propensities.

    更新日期:2020-01-04
  • Novel phosphorylation‐dependent regulation in an unstructured protein
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-01
    John F. Cannon

    This work explores how phosphorylation of an unstructured protein region in inhibitor‐2 (I2) regulates protein phosphatase‐1 (PP1) enzyme activity using molecular dynamics (MD). Free I2 is largely unstructured; however, when bound to PP1, three segments adopt a stable structure. In particular, an I2 helix (i‐helix) blocks the PP1 active site and inhibits phosphatase activity. I2 phosphorylation in the PP1‐I2 complex activates phosphatase activity without I2 dissociation. The I2 Thr74 regulatory phosphorylation site is in an unstructured domain in PP1‐I2. PP1‐I2 MD demonstrated that I2 phosphorylation promotes early steps of PP1‐I2 activation in explicit solvent models. Moreover, phosphorylation‐dependent activation occurred in PP1‐I2 complexes derived from I2 orthologs with diverse sequences from human, yeast, worm, and protozoa. This system allowed exploration of features of the 73‐residue unstructured human I2 domain critical for phosphorylation‐dependent activation. These studies revealed that components of I2 unstructured domain are strategically positioned for phosphorylation responsiveness including a transient α‐helix. There was no evidence that electrostatic interactions of I2 phosphothreonine74 influenced PP1‐I2 activation. Instead, phosphorylation altered the conformation of residues around Thr74. Phosphorylation uncurled the distance between I2 residues Glu71 to Tyr76 to promote PP1‐I2 activation, whereas reduced distances reduced activation. This I2 residue Glu71 to Tyr76 distance distribution, independently from Thr74 phosphorylation, controls I2 i‐helix displacement from the PP1 active site leading to PP1‐I2 activation.

    更新日期:2020-01-04
  • Molecular dynamics study of interactions between polymorphic actin filaments and gelsolin segment‐1
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-10-01
    Myeongsang Lee; Ellen H. Kang

    The assembly of protein actin into double‐helical filaments promotes many eukaryotic cellular processes that are regulated by actin‐binding proteins (ABPs). Actin filaments can adopt multiple conformations, known as structural polymorphism, which possibly influences the interaction between filaments and ABPs. Gelsolin is a Ca2+‐regulated ABP that severs and caps actin filaments. Gelsolin binding modulates filament structure; however, it is not known how polymorphic actin filament structures influence an interaction of gelsolin S1 with the barbed‐end of filament. Herein, we investigated how polymorphic structures of actin filaments affect the interactions near interfaces between the gelsolin segment 1 (S1) domain and the filament barbed‐end. Using all‐atom molecular dynamics simulations, we demonstrate that different tilted states of subunits modulate gelsolin S1 interactions with the barbed‐end of polymorphic filaments. Hydrogen bonding and interaction energy at the filament‐gelsolin S1 interface indicate distinct conformations of filament barbed ends, resulting in different interactions of gelsolin S1. This study demonstrates that filament's structural multiplicity plays important roles in the interactions of actin with ABPs.

    更新日期:2020-01-04
  • Prediction of the initial folding sites and the entire folding processes for Ig‐like beta‐sandwich proteins
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-30
    Panyavut Aumpuchin; Shoya Hamaue; Takeshi Kikuchi

    Describing the whole story of protein folding is currently the main enigmatic problem in molecular bioinformatics study. Protein folding mechanisms have been intensively investigated with experimental as well as simulation techniques. Since a protein folds into its specific 3D structure from a unique amino acid sequence, it is interesting to extract as much information as possible from the amino acid sequence of a protein. Analyses based on inter‐residue average distance statistics and a coarse‐grained Gō‐model simulation were conducted on Ig and FN3 domains of a titin protein to decode the folding mechanisms from their sequence data and native structure data, respectively. The central region of all domains was predicted to be an initial folding unit, that is, stable in an early state of folding. This common feature coincides well with the experimental results and underscores the significance of the β‐sandwich proteins' common structure, namely, the key strands for folding and the Greek‐key motif, which is located in the central region. We confirmed that our sequence‐based techniques were able to predict the initial folding event just next to the denatured state and that a 3D‐based Gō‐model simulation can be used to investigate the whole process of protein folding.

    更新日期:2019-12-30
  • Finding the generalized molecular principles of protein thermal stability
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-30
    Suman Hait; Saurav Mallik; Sudipto Basu; Sudip Kundu

    Are there any generalized molecular principles of thermal adaptation? Here, integrating the concepts of structural bioinformatics, sequence analysis, and classical knot theory, we develop a robust computational framework that seeks for mechanisms of thermal adaptation by comparing orthologous mesophilic‐thermophilic and mesophilic‐hyperthermophilic proteins of remarkable structural and topological similarities, and still leads us to context‐independent results. A comprehensive analysis of 4741 high‐resolution, non‐redundant X‐ray crystallographic structures collected from 11 hyperthermophilic, 32 thermophilic and 53 mesophilic prokaryotes unravels at least five “nearly universal” signatures of thermal adaptation, irrespective of the enormous sequence, structure, and functional diversity of the proteins compared. A careful investigation further extracts a set of amino acid changes that can potentially enhance protein thermal stability, and remarkably, these mutations are overrepresented in protein crystallization experiments, in disorder‐to‐order transitions and in engineered thermostable variants of existing mesophilic proteins. These results could be helpful to find a precise, global picture of thermal adaptation.

    更新日期:2019-12-30
  • SAXSDom: Modeling multidomain protein structures using small‐angle X‐ray scattering data
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-27
    Jie Hou; Badri Adhikari; John J. Tanner; Jianlin Cheng

    Many proteins are composed of several domains that pack together into a complex tertiary structure. Multidomain proteins can be challenging for protein structure modeling, particularly those for which templates can be found for individual domains but not for the entire sequence. In such cases, homology modeling can generate high quality models of the domains but not for the orientations between domains. Small‐angle X‐ray scattering (SAXS) reports the structural properties of entire proteins and has the potential for guiding homology modeling of multidomain proteins. In this article, we describe a novel multidomain protein assembly modeling method, SAXSDom that integrates experimental knowledge from SAXS with probabilistic Input‐Output Hidden Markov model to assemble the structures of individual domains together. Four SAXS‐based scoring functions were developed and tested, and the method was evaluated on multidomain proteins from two public datasets. Incorporation of SAXS information improved the accuracy of domain assembly for 40 out of 46 critical assessment of protein structure prediction multidomain protein targets and 45 out of 73 multidomain protein targets from the ab initio domain assembly dataset. The results demonstrate that SAXS data can provide useful information to improve the accuracy of domain‐domain assembly. The source code and tool packages are available at https://github.com/jianlin-cheng/SAXSDom.

    更新日期:2019-12-27
  • Structural and functional characterization of Solanum tuberosum VDAC36
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-20
    Maximilien Lopes‐Rodrigues; André Matagne; David Zanuy; Carlos Alemán; Eric A. Perpète; Catherine Michaux

    As it forms water‐filled channel in the mitochondria outer membrane and diffuses essential metabolites such as NADH and ATP, the voltage‐dependent anion channel (VDAC) protein family plays a central role in all eukaryotic cells. In comparison with their mammalian homologues, little is known about the structural and functional properties of plant VDACs. In the present contribution, one of the two VDACs isoforms of Solanum tuberosum, stVDAC36, has been successfully overexpressed and refolded by an in‐house method, as demonstrated by the information on its secondary and tertiary structure gathered from circular dichroism and intrinsic fluorescence. Cross‐linking and molecular modeling studies have evidenced the presence of dimers and tetramers, and they suggest the formation of an intermolecular disulfide bond between two stVDAC36 monomers. The pore‐forming activity was also assessed by liposome swelling assays, indicating a typical pore diameter between 2.0 and 2.7 nm. Finally, insights about the ATP binding inside the pore are given by docking studies and electrostatic calculations.

    更新日期:2019-12-20
  • The crystal structure of the mycobacterial trehalose monomycolate transport factor A, TtfA, reveals an atypical fold
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-19
    Kien Lam Ung; Husam M. A. B. Alsarraf; Laurent Kremer; Mickaël Blaise

    Trehalose monomycolate (TMM) represents an essential element of the mycobacterial envelope. While synthesized in the cytoplasm, TMM is transported across the inner membrane by MmpL3 but, little is known regarding the MmpL3 partners involved in this process. Recently, the TMM transport factor A (TtfA) was found to form a complex with MmpL3 and to participate in TMM transport, although its biological role remains to be established. Herein, we report the crystal structure of the Mycobacterium smegmatis TtfA core domain. The phylogenetic distribution of TtfA homologues in non‐mycolate containing bacteria suggests that TtfA may exert additional functions.

    更新日期:2019-12-19
  • Escapement mechanisms: Efficient free energy transduction by reciprocally‐coupled gating
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-13
    Charles W. Carter

    Conversion of the free energy of NTP hydrolysis efficiently into mechanical work and/or information by transducing enzymes sustains living systems far from equilibrium, and so has been of interest for many decades. Detailed molecular mechanisms, however, remain puzzling and incomplete. We previously reported that catalysis of tryptophan activation by tryptophanyl‐tRNA synthetase, TrpRS, requires relative domain motion to re‐position the catalytic Mg2+ ion, noting the analogy between that conditional hydrolysis of ATP and the escapement mechanism of a mechanical clock. The escapement allows the time‐keeping mechanism to advance discretely, one gear at a time, if and only if the pendulum swings, thereby converting energy from the weight driving the pendulum into rotation of the hands. Coupling of catalysis to domain motion, however, mimics only half of the escapement mechanism, suggesting that domain motion may also be reciprocally coupled to catalysis, completing the escapement metaphor. Computational studies of the free energy surface restraining the domain motion later confirmed that reciprocal coupling: the catalytic domain motion is thermodynamically unfavorable unless the PPi product is released from the active site. These two conditional phenomena—demonstrated together only for the TrpRS mechanism—function as reciprocally‐coupled gates. As we and others have noted, such an escapement mechanism is essential to the efficient transduction of NTP hydrolysis free energy into other useful forms of mechanical or chemical work and/or information. Some implementation of both gating mechanisms—catalysis by domain motion and domain motion by catalysis—will thus likely be found in many other systems.

    更新日期:2019-12-17
  • Coarse‐grained and atomic resolution biomolecular docking with the ATTRACT approach
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-13
    Glenn Glashagen; Sjoerd de Vries; Urszula Uciechowska‐Kaczmarzyk; Sergey A. Samsonov; Samuel Murail; Pierre Tuffery; Martin Zacharias

    The ATTRACT protein‐protein docking program has been employed to predict protein‐protein complex structures in CAPRI rounds 38‐45. For 11 out of 16 targets acceptable or better quality solutions have been submitted (~70%). It includes also several cases of peptide‐protein docking and the successful prediction of the geometry of carbohydrate‐protein interactions. The option of combining rigid body minimization and simultaneous optimization in collective degrees of freedom based on elastic network modes was employed and systematically evaluated. Application to a large benchmark set indicates a modest improvement in docking performance compared to rigid docking. Possible further improvements of the docking approach in particular at the scoring and the flexible refinement steps are discussed.

    更新日期:2019-12-13
  • Structure prediction of biological assemblies using GALAXY in CAPRI rounds 38‐45
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-12-10
    Taeyong Park, Hyeonuk Woo, Minkyung Baek, Jinsol Yang, Chaok Seok

    We participated in CARPI rounds 38‐45 both as a server predictor and a human predictor. These CAPRI rounds provided excellent opportunities for testing prediction methods for three classes of protein interactions, that is, protein‐protein, protein‐peptide, and protein‐oligosaccharide interactions. Both template‐based methods (GalaxyTBM for monomer protein, GalaxyHomomer for homo‐oligomer protein, GalaxyPepDock for protein‐peptide complex) and ab initio docking methods (GalaxyTongDock and GalaxyPPDock for protein oligomer, GalaxyPepDock‐ab‐initio for protein‐peptide complex, GalaxyDock2 and Galaxy7TM for protein‐oligosaccharide complex) have been tested. Template‐based methods depend heavily on the availability of proper templates and template‐target similarity, and template‐target difference is responsible for inaccuracy of template‐based models. Inaccurate template‐based models could be improved by our structure refinement and loop modeling methods based on physics‐based energy optimization (GalaxyRefineComplex and GalaxyLoop) for several CAPRI targets. Current ab initio docking methods require accurate protein structures as input. Small conformational changes from input structure could be accounted for by our docking methods, producing one of the best models for several CAPRI targets. However, predicting large conformational changes involving protein backbone is still challenging, and full exploration of physics‐based methods for such problems is still to come.

    更新日期:2019-12-11
  • The effects of somatic mutations on EGFR interaction with anti‐EGFR monoclonal antibodies: Implication for acquired resistance
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-02
    Maryam Tabasinezhad, Eskanadr Omidinia, Yeganeh Talebkhan, Mir Davood Omrani, Fereidoun Mahboudi, Hamid Ghaedi, Wolfgang Wenzel

    A number of mutations in the epidermal growth factor receptor (EGFR) have been identified that imparts resistance to anti‐EGFR monoclonal antibodies (mAbs) in clinical and preclinical samples. Primary or acquired resistance to targeted therapy will eventually limit the clinical benefit of anticancer mAbs. The aim of the current study was to perform computational analysis to investigate the structural implications of the EGFR somatic mutations on its complexes with the four anti‐EGFR mAbs (Cetuximab, Panitumumab, Necitumumab, and Matuzumab). Docking analysis and molecular dynamics (MD) simulations were performed to understand the plausible structural and dynamical implications caused by somatic mutations available in the Catalogue of Somatic Mutations in Cancer database on the EGFR and anti‐EGFR mAbs. We found that EGFRS492R and EGFRV441I in complex with Cetuximab, EGFRR377S and EGFRS447Y in complex with Panitumumab, and EGFRV441I in complex with Necitumumab have a weakest binding affinity in comparison to EGFRWT in complex with the relevant mAb. Taken together with the results obtained from docking analysis and MD simulations, the present findings may suggest that, the S492R and V441I mutations confer resistance to Cetuximab, R377S and S447Y mutations mediate resistance to Panitumumab and finally, V441I mutation also confers resistance to Necitumumab.

    更新日期:2019-12-09
  • Enabling full‐length evolutionary profiles based deep convolutional neural network for predicting DNA‐binding proteins from sequence
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-08
    Sucheta Chauhan, Shandar Ahmad

    Sequence based DNA‐binding protein (DBP) prediction is a widely studied biological problem. Sliding windows on position specific substitution matrices (PSSMs) rows predict DNA‐binding residues well on known DBPs but the same models cannot be applied to unequally sized protein sequences. PSSM summaries representing column averages and their amino‐acid wise versions have been effectively used for the task, but it remains unclear if these features carry all the PSSM's predictive power, traditionally harnessed for binding site predictions. Here we evaluate if PSSMs scaled up to a fixed size by zero‐vector padding (pPSSM) could perform better than the summary based features on similar models. Using multilayer perceptron (MLP) and deep convolutional neural network (CNN), we found that (a) Summary features work well for single‐genome (human‐only) data but are outperformed by pPSSM for diverse PDB‐derived data sets, suggesting greater summary‐level redundancy in the former, (b) even when summary features work comparably well with pPSSM, a consensus on the two outperforms both of them (c) CNN models comprehensively outperform their corresponding MLP models and (d) actual predicted scores from different models depend on the choice of input feature sets used whereas overall performance levels are model‐dependent in which CNN leads the accuracy.

    更新日期:2019-12-09
  • The 3A6‐TCR/superagonist/HLA‐DR2a complex shows similar interface and reduced flexibility compared to the complex with self‐peptide
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-06-25
    Ilaria Salutari, Roland Martin, Amedeo Caflisch

    T‐cell receptor (TCR) recognition of the myelin basic protein (MBP) peptide presented by major histocompatibility complex (MHC) protein HLA‐DR2a, one of the MHC class II alleles associated with multiple sclerosis, is highly variable. Interactions in the trimolecular complex between the TCR of the MBP83‐99‐specific T cell clone 3A6 with the MBP‐peptide/HLA‐DR2a (abbreviated TCR/pMHC) lead to substantially different proliferative responses when comparing the wild‐type decapeptide MBP90‐99 and a superagonist peptide, which differs mainly in the residues that point toward the TCR. Here, we investigate the influence of the peptide sequence on the interface and intrinsic plasticity of the TCR/pMHC trimolecular and pMHC bimolecular complexes by molecular dynamics simulations. The intermolecular contacts at the TCR/pMHC interface are similar for the complexes with the superagonist and the MBP self‐peptide. The orientation angle between TCR and pMHC fluctuates less in the complex with the superagonist peptide. Thus, the higher structural stability of the TCR/pMHC tripartite complex with the superagonist peptide, rather than a major difference in binding mode with respect to the self‐peptide, seems to be responsible for the stronger proliferative response.

    更新日期:2019-12-09
  • Structural characterization of β‐ketoacyl ACP synthase I bound to platencin and fragment screening molecules at two substrate binding sites
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-17
    Edward I. Patterson, Jeffrey D. Nanson, Jan Abendroth, Cassie Bryan, Banumathi Sankaran, Peter J. Myler, Jade K. Forwood

    The bacterial fatty acid pathway is essential for membrane synthesis and a range of other metabolic and cellular functions. The β‐ketoacyl‐ACP synthases carry out the initial elongation reaction of this pathway, utilizing acetyl‐CoA as a primer to elongate malonyl‐ACP by two carbons, and subsequent elongation of the fatty acyl‐ACP substrate by two carbons. Here we describe the structures of the β‐ketoacyl‐ACP synthase I from Brucella melitensis in complex with platencin, 7‐hydroxycoumarin, and (5‐thiophen‐2‐ylisoxazol‐3‐yl)methanol. The enzyme is a dimer and based on structural and sequence conservation, harbors the same active site configuration as other β‐ketoacyl‐ACP synthases. The platencin binding site overlaps with the fatty acyl compound supplied by ACP, while 7‐hydroxyl‐coumarin and (5‐thiophen‐2‐ylisoxazol‐3‐yl)methanol bind at the secondary fatty acyl binding site. These high‐resolution structures, ranging between 1.25 and 1.70 å resolution, provide a basis for in silico inhibitor screening and optimization, and can aid in rational drug design by revealing the high‐resolution binding interfaces of molecules at the malonyl‐ACP and acyl‐ACP active sites.

    更新日期:2019-12-09
  • Multimapping confounds ribosome profiling analysis: A case‐study of the Hsp90 molecular chaperone
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-19
    Jackson C. Halpin, Radhika Jangi, Timothy O. Street

    Ribosome profiling (Ribo‐seq) can potentially provide detailed information about ribosome position on transcripts and estimates of protein translation levels in vivo. Hsp90 chaperones, which play a critical role in stress tolerance, have characteristic patterns of differential expression under nonstressed and heat shock conditions. By analyzing published Ribo‐seq data for the Hsp90 chaperones in S. cerevisiae, we find wide‐ranging artifacts originating from “multimapping” reads (reads that cannot be uniquely assigned to one position), which constitute ~25% of typical S. cerevisiae Ribo‐seq datasets and ~80% of the reads from HEK293 cells. Estimates of Hsp90 protein production as determined by Ribo‐seq are reproducible but not robust, with inferred expression levels that can change 10‐fold depending on how multimapping reads are processed. The differential expression of Hsp90 chaperones under nonstressed and heat shock conditions creates artificial peaks and valleys in their ribosome profiles that give a false impression of regulated translational pausing. Indeed, we find that multimapping can even create an appearance of reproducibility to the shape of the Hsp90 ribosome profiles from biological replicates. Adding further complexity, this artificial reproducibility is dependent on the computational method used to construct the ribosome profile. Given the ubiquity of multimapping reads in Ribo‐seq experiments and the complexity of artifacts associated with multimapping, we developed a publicly available computational tool to identify transcripts most at risk for multimapping artifacts. In doing so, we identify biological pathways that are enriched in multimapping transcripts, meaning that particular biological pathways will be highly susceptible to multimapping artifacts.

    更新日期:2019-12-09
  • Structural basis for −35 element recognition by σ4 chimera proteins and their interactions with PmrA response regulator
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-22
    Yuan‐Chao Lou, Chun‐Chi Chou, Hsin‐Hong Yeh, Chia‐Yu Chien, Sushant Sadotra, Chun‐Hua Hsu, Chinpan Chen

    In class II transcription activation, the transcription factor normally binds to the promoter near the −35 position and contacts the domain 4 of σ factors (σ4) to activate transcription. However, σ4 of σ70 appears to be poorly folded on its own. Here, by fusing σ4 with the RNA polymerase β‐flap‐tip‐helix, we constructed two σ4 chimera proteins, one from σ70 and another from σS of Klebsiella pneumoniae. The two chimera proteins well folded into a monomeric form with strong binding affinities for −35 element DNA. Determining the crystal structure of in complex with −35 element DNA revealed that adopts a similar structure as σ4 in the Escherichia coli RNA polymerase σS holoenzyme and recognizes −35 element DNA specifically by several conserved residues from the helix‐turn‐helix motif. By using nuclear magnetic resonance (NMR), was demonstrated to recognize −35 element DNA similar to . Carr‐Purcell‐Meiboom‐Gill relaxation dispersion analyses showed that the N‐terminal helix and the β‐flap‐tip‐helix of have a concurrent transient α‐helical structure and DNA binding reduced the slow dynamics on . Finally, only was shown to interact with the response regulator PmrA and its promoter DNA. The chimera proteins are capable of −35 element DNA recognition and can be used for study with transcription factors or other factors that interact with domain 4 of σ factors.

    更新日期:2019-12-09
  • An NMR and MD study of complexes of bacteriophage lambda lysozyme with tetra‐ and hexa‐N‐acetylchitohexaose
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-26
    Aysegul Turupcu, Alice M. Bowen, Alexandre Di Paolo, André Matagne, Chris Oostenbrink, Christina Redfield, Lorna J. Smith

    The X‐ray structure of lysozyme from bacteriophage lambda (λ lysozyme) in complex with the inhibitor hexa‐N‐acetylchitohexaose (NAG6) (PDB: 3D3D) has been reported previously showing sugar units from two molecules of NAG6 bound in the active site. One NAG6 is bound with four sugar units in the ABCD sites and the other with two sugar units in the E′F′ sites potentially representing the cleavage reaction products; each NAG6 cross links two neighboring λ lysozyme molecules. Here we use NMR and MD simulations to study the interaction of λ lysozyme with the inhibitors NAG4 and NAG6 in solution. This allows us to study the interactions within the complex prior to cleavage of the polysaccharide. 1HN and 15N chemical shifts of λ lysozyme resonances were followed during NAG4/NAG6 titrations. The chemical shift changes were similar in the two titrations, consistent with sugars binding to the cleft between the upper and lower domains; the NMR data show no evidence for simultaneous binding of a NAG6 to two λ lysozyme molecules. Six 150 ns MD simulations of λ lysozyme in complex with NAG4 or NAG6 were performed starting from different conformations. The simulations with both NAG4 and NAG6 show stable binding of sugars across the D/E active site providing low energy models for the enzyme‐inhibitor complexes. The MD simulations identify different binding subsites for the 5th and 6th sugars consistent with the NMR data. The structural information gained from the NMR experiments and MD simulations have been used to model the enzyme‐peptidoglycan complex.

    更新日期:2019-12-09
  • Ligand nanovectorization using graphene to target cellular death receptors of cancer cell
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-29
    Nicolas Arroyo, Guillaume Herlem, Fabien Picaud

    Tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL) is nowadays envisaged as a natural cytokine useful in nanomedicine to eradicate the cancer cells and not the healthy surrounding ones. However, it suffers from cell resistance and strong dispersion in body to prove its efficiency. The understanding at the molecular level of the TRAIL interaction with death receptors (DRs) on cancer cells is thus of fundamental importance to improve its action. We demonstrate here via molecular simulations that TRAIL can bind to its both agonistic DRs (ie, DR4 and DR5) with a preference for DR4. In this study, the role of a graphene nanoflake as a potential cargo for TRAIL is examined. Furthermore, both TRAIL self‐assembling and TRAIL affinity when adsorbed on graphene are considered to enhance efficacy toward the targeted cancer cell. Our modelization results show that TRAIL can bind to DR4 and DR5 when transported by graphene nanoflake, as a proof of concept.

    更新日期:2019-12-09
  • The structure of the extended E2 DNA‐binding domain of the bovine papillomavirus‐1
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-01
    Ludmila Leroy, João Alexandre Ribeiro Gonçalves Barbosa, Gonzalo de Prat‐Gay, Igor Polikarpov, Carlos Basílio Pinheiro

    Bovine papillomavirus proteins were extensively studied as a prototype for the human papillomavirus. Here, the crystal structure of the extended E2 DNA‐binding domain of the dominant transcription regulator from the bovine papillomavirus strain 1 is described in the space group P3121. We found two protein functional dimers packed in the asymmetric unit. This new protein arrangement inside the crystal led to the reduction of the mobility of a previously unobserved loop directly involved in the protein‐DNA interaction, which was then modeled for the first time.

    更新日期:2019-12-09
  • F508del disturbs the dynamics of the nucleotide binding domains of CFTR before and after ATP hydrolysis
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-17
    Bárbara Abreu, Emanuel F. Lopes, A. S. F. Oliveira, Cláudio M. Soares

    The cystic fibrosis transmembrane conductance regulator (CFTR) channel is an ion channel responsible for chloride transport in epithelia and it belongs to the class of ABC transporters. The deletion of phenylalanine 508 (F508del) in CFTR is the most common mutation responsible for cystic fibrosis. Little is known about the effect of the mutation in the isolated nucleotide binding domains (NBDs), on dimer dynamics, ATP hydrolysis and even on nucleotide binding. Using molecular dynamics simulations of the human CFTR NBD dimer, we showed that F508del increases, in the prehydrolysis state, the inter‐motif distance in both ATP binding sites (ABP) when ATP is bound. Additionally, a decrease in the number of catalytically competent conformations was observed in the presence of F508del. We used the subtraction technique to study the first 300 ps after ATP hydrolysis in the catalytic competent site and found that the F508del dimer evidences lower conformational changes than the wild type. Using longer simulation times, the magnitude of the conformational changes in both forms increases. Nonetheless, the F508del dimer shows lower C‐α RMS values in comparison to the wild‐type, on the F508del loop, on the residues surrounding the catalytic site and the portion of NBD2 adjacent to ABP1. These results provide evidence that F508del interferes with the NBD dynamics before and after ATP hydrolysis. These findings shed a new light on the effect of F508del on NBD dynamics and reveal a novel mechanism for the influence of F508del on CFTR.

    更新日期:2019-12-09
  • A free‐energy landscape for the glucagon‐like peptide 1 receptor GLP1R
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-02
    Raphael Alhadeff, Arieh Warshel

    G‐protein‐coupled receptors (GPCRs) are among the most important receptors in human physiology and pathology. They serve as master regulators of numerous key processes and are involved in as well as cause debilitating diseases. Consequently, GPCRs are among the most attractive targets for drug design and pharmaceutical interventions (>30% of drugs on the market). The glucagon‐like peptide 1 (GLP‐1) hormone receptor GLP1R is closely involved in insulin secretion by pancreatic β‐cells and constitutes a major druggable target for the development of anti‐diabetes and obesity agents. GLP1R structure was recently solved, with ligands, allosteric modulators and as part of a complex with its cognate G protein. However, the translation of this structural data into structure/function understanding remains limited. The current study functionally characterizes GLP1R with special emphasis on ligand and cellular partner binding interactions and presents a free‐energy landscape as well as a functional model of the activation cycle of GLP1R. Our results should facilitate a deeper understanding of the molecular mechanism underlying GLP1R activation, forming a basis for improved development of targeted therapeutics for diabetes and related disorders.

    更新日期:2019-12-09
  • Discovery of receptor‐ligand interfaces in the immunoglobulin superfamily
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-29
    Nelson Gil, Eduardo J. Fajardo, Andras Fiser

    Cell‐surface‐anchored immunoglobulin superfamily (IgSF) proteins are widespread throughout the human proteome, forming crucial components of diverse biological processes including immunity, cell‐cell adhesion, and carcinogenesis. IgSF proteins generally function through protein‐protein interactions carried out between extracellular, membrane‐bound proteins on adjacent cells, known as trans‐binding interfaces. These protein‐protein interactions constitute a class of pharmaceutical targets important in the treatment of autoimmune diseases, chronic infections, and cancer. A molecular‐level understanding of IgSF protein‐protein interactions would greatly benefit further drug development. A critical step toward this goal is the reliable identification of IgSF trans‐binding interfaces. We propose a novel combination of structure and sequence information to identify trans‐binding interfaces in IgSF proteins. We developed a structure‐based binding interface prediction approach that can identify broad regions of the protein surface that encompass the binding interfaces and suggests that IgSF proteins possess binding supersites. These interfaces could theoretically be pinpointed using sequence‐based conservation analysis, with performance approaching the theoretical upper limit of binding interface prediction accuracy, but achieving this in practice is limited by the current ability to identify an appropriate multiple sequence alignment for conservation analysis. However, an important contribution of combining the two orthogonal methods is that agreement between these approaches can estimate the reliability of the predictions. This approach was benchmarked on the set of 22 IgSF proteins with experimentally solved structures in complex with their ligands. Additionally, we provide structure‐based predictions and reliability scores for the 62 IgSF proteins with known structure but yet uncharacterized binding interfaces.

    更新日期:2019-12-09
  • A deep dense inception network for protein beta‐turn prediction
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-23
    Chao Fang, Yi Shang, Dong Xu

    Beta‐turn prediction is useful in protein function studies and experimental design. Although recent approaches using machine‐learning techniques such as support vector machine (SVM), neural networks, and K nearest neighbor have achieved good results for beta‐turn prediction, there is still significant room for improvement. As previous predictors utilized features in a sliding window of 4‐20 residues to capture interactions among sequentially neighboring residues, such feature engineering may result in incomplete or biased features and neglect interactions among long‐range residues. Deep neural networks provide a new opportunity to address these issues. Here, we proposed a deep dense inception network (DeepDIN) for beta‐turn prediction, which takes advantage of the state‐of‐the‐art deep neural network design of dense networks and inception networks. A test on a recent BT6376 benchmark data set shows that DeepDIN outperformed the previous best tool BetaTPred3 significantly in both the overall prediction accuracy and the nine‐type beta‐turn classification accuracy. A tool, called MUFold‐BetaTurn, was developed, which is the first beta‐turn prediction tool utilizing deep neural networks. The tool can be downloaded at http://dslsrv8.cs.missouri.edu/~cf797/MUFoldBetaTurn/download.html.

    更新日期:2019-12-09
  • DRoP: Automated detection of conserved solvent‐binding sites on proteins
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-23
    Bradley M. Kearney, Michael Schwabe, Kendra C. Marcus, Daniel M. Roberts, Michelle Dechene, Paul Swartz, Carla Mattos

    Water and ligand binding play critical roles in the structure and function of proteins, yet their binding sites and significance are difficult to predict a priori. Multiple solvent crystal structures (MSCS) is a method where several X‐ray crystal structures are solved, each in a unique solvent environment, with organic molecules that serve as probes of the protein surface for sites evolved to bind ligands, while the first hydration shell is essentially maintained. When superimposed, these structures contain a vast amount of information regarding hot spots of protein‐protein or protein‐ligand interactions, as well as conserved water‐binding sites retained with the change in solvent properties. Optimized mining of this information requires reliable structural data and a consistent, objective analysis tool. Detection of related solvent positions (DRoP) was developed to automatically organize and rank the water or small organic molecule binding sites within a given set of structures. It is a flexible tool that can also be used in conserved water analysis given multiple structures of any protein independent of the MSCS method. The DRoP output is an HTML format list of the solvent sites ordered by conservation rank in its population within the set of structures, along with renumbered and recolored PDB files for visualization and facile analysis. Here, we present a previously unpublished set of MSCS structures of bovine pancreatic ribonuclease A (RNase A) and use it together with published structures to illustrate the capabilities of DRoP.

    更新日期:2019-12-09
  • Modification of lactoferrin by peroxynitrite reduces its antibacterial activity and changes protein structure
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-23
    Amani Y. Alhalwani, Rachel L. Davey, Navneeta Kaul, Scott A. Barbee, J. Alex Huffman

    Lactoferrin (LF) is a multifunctional protein that plays important physiological roles as one of the most concentrated proteins in many human and other mammalian fluids and tissues. In particular, LF provides antibacterial properties to human milk, saliva, and tear fluid. LF also protects against stress‐induced lipid peroxidation at inflammation sites through its iron‐binding ability. Previous studies have shown that LF can be efficiently nitrated via biologically relevant mediators such as peroxynitrite (ONOO−), which are also present at high intracellular concentrations during inflammation and nitrosative stress. Here, we examine changes in antibacterial properties and structure of LF following ONOO− treatment. The reaction induces nitration of tyrosine and tryptophan residues, which are commonly used as biomarker molecules for several diseases. Treatment with ONOO− at a 10/1 M ratio of ONOO− to tyrosine inhibited all antibacterial activity exhibited by native LF. Secondary structural changes in LF were assessed using circular dichroism spectroscopy. Nitration products with and without the addition of Fe3+ show significant reduction in alpha‐helical properties, suggesting partial protein unfolding. Iron‐binding capacity of LF was also reduced after treatment with ONOO−, suggesting a decreased ability of LF to protect against cellular damage. LC‐MS/MS spectrometry was used to identify LF peptide fragments nitrated by ONOO−, including tyrosine residue Y92 located in the iron‐binding region. These results suggest that posttranslational modification of LF by ONOO− could be an important pathway to exacerbate infection, for example, in inflamed tissues and to reduce the ability of LF to act as an immune responder and decrease oxidative damage.

    更新日期:2019-12-09
  • Antimicrobial activity and structure of a consensus human β‐defensin and its comparison to a novel putative hBD10
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-30
    Alexis Rodriguez, Marie Ø. Pedersen, Elba Villegas, Bruno Rivas‐Santiago, Jessica Villegas‐Moreno, Carlos Amero, Raymond S. Norton, Gerardo Corzo

    The spread of multidrug resistant bacteria owing to the intensive use of antibiotics is challenging current antibiotic therapies, and making the discovery and evaluation of new antimicrobial agents a high priority. The evaluation of novel peptide sequences of predicted antimicrobial peptides from different sources is valuable approach to identify alternative antibiotic leads. Two strategies were pursued in this study to evaluate novel antimicrobial peptides from the human β‐defensin family (hBD). In the first, a 32‐residue peptide was designed based on the alignment of all available hBD primary structures, while in the second a putative 35‐residue peptide, hBD10, was mined from the gene DEFB110. Both hBDconsensus and hBD10 were chemically synthesized, folded and purified. They showed antimicrobial activity against Escherichia coli, Staphylococcus aureus, and Mycobacterium tuberculosis, but were not hemolytic on human red blood cells. The NMR‐based solution structure of hBDconsensus revealed that it adopts a classical β‐defensin fold and disulfide connectivities. Even though the mass spectrum of hBD10 confirmed the formation of three disulfide bonds, it showed limited dispersion in 1H NMR spectra and structural studies were not pursued. The evaluation of different β‐defensin structures may identify new antimicrobial agents effective against multidrug‐resistant bacterial strains.

    更新日期:2019-12-09
  • Design of a basigin‐mimicking inhibitor targeting the malaria invasion protein RH5
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-02
    Shira Warszawski, Elya Dekel, Ivan Campeotto, Jennifer M. Marshall, Katherine E. Wright, Oliver Lyth, Orli Knop, Neta Regev‐Rudzki, Matthew K. Higgins, Simon J. Draper, Jake Baum, Sarel J. Fleishman

    Many human pathogens use host cell‐surface receptors to attach and invade cells. Often, the host‐pathogen interaction affinity is low, presenting opportunities to block invasion using a soluble, high‐affinity mimic of the host protein. The Plasmodium falciparum reticulocyte‐binding protein homolog 5 (RH5) provides an exciting candidate for mimicry: it is highly conserved and its moderate affinity binding to the human receptor basigin (KD ≥1 μM) is an essential step in erythrocyte invasion by this malaria parasite. We used deep mutational scanning of a soluble fragment of human basigin to systematically characterize point mutations that enhance basigin affinity for RH5 and then used Rosetta to design a variant within the sequence space of affinity‐enhancing mutations. The resulting seven‐mutation design exhibited 1900‐fold higher affinity (KD approximately 1 nM) for RH5 with a very slow binding off rate (0.23 h−1) and reduced the effective Plasmodium growth‐inhibitory concentration by at least 10‐fold compared to human basigin. The design provides a favorable starting point for engineering on‐rate improvements that are likely to be essential to reach therapeutically effective growth inhibition.

    更新日期:2019-12-09
  • Transcriptional regulatory module analysis reveals that bridge proteins reconcile multiple signals in extracellular electron transfer pathways
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-25
    Dewu Ding, Chuanjun Shu, Xiao Sun

    Shewanella oneidensis MR‐1 shows remarkable respiratory versatility with a large variety of extracellular electron acceptors (termed extracellular electron transfer, EET). To utilize the various electron acceptors, the bacterium must employ complex regulatory mechanisms to elicit the relevant EET pathways. To investigate the relevant mechanisms, we integrated EET genes and related transcriptional factors (TFs) into transcriptional regulatory modules (TRMs) and showed that many bridge proteins in these modules were signal proteins, which generally contained one or more signal processing domains (eg, GGDEF, EAL, PAS, etc.). Since Shewanella has to respond to diverse environmental conditions despite encoding few EET‐relevant TFs, the overabundant signal proteins involved in the TRMs can help decipher the mechanism by which these microbes elicit a wide range of condition‐specific responses. By combining proteomic data and protein bioinformatic analysis, we demonstrated that diverse signal proteins reconciled the different EET pathways, and we discussed the functional roles of signal proteins involved in the well‐known MtrCAB pathway. Additionally, we showed that the signal proteins SO_2145 and SO_1417 played central roles in triggering EET pathways in anaerobic environments. Taken together, our results suggest that signal proteins have a profound impact on the transcriptional regulation of EET genes and thus have potential applications in microbial fuel cells.

    更新日期:2019-12-09
  • Comparison of Rosetta flexible‐backbone computational protein design methods on binding interactions
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-10
    Amanda L. Loshbaugh, Tanja Kortemme

    Computational design of binding sites in proteins remains difficult, in part due to limitations in our current ability to sample backbone conformations that enable precise and accurate geometric positioning of side chains during sequence design. Here we present a benchmark framework for comparison between flexible‐backbone design methods applied to binding interactions. We quantify the ability of different flexible backbone design methods in the widely used protein design software Rosetta to recapitulate observed protein sequence profiles assumed to represent functional protein/protein and protein/small molecule binding interactions. The CoupledMoves method, which combines backbone flexibility and sequence exploration into a single acceptance step during the sampling trajectory, better recapitulates observed sequence profiles than the BackrubEnsemble and FastDesign methods, which separate backbone flexibility and sequence design into separate acceptance steps during the sampling trajectory. Flexible‐backbone design with the CoupledMoves method is a powerful strategy for reducing sequence space to generate targeted libraries for experimental screening and selection.

    更新日期:2019-12-09
  • In vitro and in silico studies on membrane interactions of diverse Capsicum annuum flower γ‐thionin peptides
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-19
    Siddhanta Nikte, Apurva Gahankari, Javed Mulla, Durba Sengupta, Manali Joshi, Vaijayanti Tamhane

    Thionins are small, cysteine‐rich peptides that play an important role in plant defense, primarily through their interactions with membranes. Eight novel γ‐thionin peptides (CanThio1‐8) were isolated from the flower of Capsicum annuum. Sequence analysis revealed that the peptides cluster into three groups. A representative peptide from each group (CanThio1, 2, and 3) was used for experimental characterization. Interestingly, peptides were found to possess some cytotoxic activity against normal human embryonic kidney cell line but higher cytotoxicity against cancer cell line MCF‐7. CanThio3 peptide was chosen as a representative peptide to study the molecular mechanism of action on membranes. Microsecond timescale atomistic simulations of CanThio3 were performed in the presence of a POPC (1‐palmitoyl‐2‐oleoyl‐sn‐glycero‐3‐phosphocholine) lipid bilayer. Simulations revealed that CanThio3 interacts with the bilayer and causes lipid thinning in the vicinity. Nonpolar amino acids specific to the α‐core region of CanThio3 along with nonpolar residues in the γ‐core region are seen to interact with the lipid tails. The differences in the amino acid sequence of CanThio peptides in these regions explain the variability in cytotoxic activities. In summary, our results demonstrate the membrane‐mediated activity of a novel series of γ‐thionin peptides from C. annuum.

    更新日期:2019-12-09
  • Solution NMR structure of CGL2373, a polyketide cyclase‐like protein from Corynebacterium glutamicum
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-07-17
    Cong Cai, Yao Nie, Yixuan Gong, Shuangli Li, Theresa A. Ramelot, Michael A. Kennedy, Xiali Yue, Jiang Zhu, Maili Liu, Yunhuang Yang

    Protein CGL2373 from Corynebacterium glutamicum was previously proposed to be a member of the polyketide_cyc2 family, based on amino‐acid sequence and secondary structure features derived from NMR chemical shift assignments. We report here the solution NMR structure of CGL2373, which contains three α‐helices and one antiparallel β‐sheet and adopts a helix‐grip fold. This structure shows moderate similarities to the representative polyketide cyclases, TcmN, WhiE, and ZhuI. Nevertheless, unlike the structures of these homologs, CGL2373 structure looks like a half‐open shell with a much larger pocket, and key residues in the representative polyketide cyclases for binding substrate and catalyzing aromatic ring formation are replaced with different residues in CGL2373. Also, the gene cluster where the CGL2373‐encoding gene is located in C. glutamicum contains additional genes encoding nucleoside diphosphate kinase, folylpolyglutamate synthase, and valine‐tRNA ligase, different from the typical gene cluster encoding polyketide cyclase in Streptomyces. Thus, although CGL2373 is structurally a polyketide cyclase‐like protein, the function of CGL2373 may differ from the known polyketide cyclases and needs to be further investigated. The solution structure of CGL2373 lays a foundation for in silico ligand screening and binding site identifying in future functional study.

    更新日期:2019-12-09
  • Nuclear magnetic resonance solution structure of Pisum sativum defensin 2 provides evidence for the presence of hydrophobic surface‐clusters
    Proteins Struct. Funct. Bioinform. (IF 2.501) Pub Date : 2019-08-05
    Ramon Pinheiro‐Aguiar, Virginia S. G. do Amaral, Iuri B. Pereira, Eleonora Kurtenbach, Fabio C. L. Almeida

    Pisum sativum defensin 2 (Psd2) is a small (4.7 kDa) antifungal peptide whose structure is held together by four conserved disulfide bridges. Psd2 shares the cysteine‐stabilized alpha‐beta (CSαβ) fold, which lacks a regular hydrophobic core. All hydrophobic residues are exposed to the surface, except for leucine 6. They are clustered in the surface formed by two loops, between β1 and α‐helix and β2 and β3 sheets. The observation of surface hydrophobic clusters reveals a remarkable evolution of the CSαβ fold to expose and reorganize hydrophobic residues, which facilitates creating versatile binding sites.

    更新日期:2019-12-09
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