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  • 更新日期:2018-07-16
  • Sesquiterpene Synthase‐Catalysed Formation of a New Medium‐Sized Cyclic Terpenoid Ether from Farnesyl Diphosphate Analogues
    ChemBioChem (IF 2.774) Pub Date : 2018-05-26
    Florence Huynh; Daniel J. Grundy; Robert L. Jenkins; David J. Miller; Rudolf K. Allemann
    更新日期:2018-07-16
  • Three S's for Aptamer‐Mediated Control of DNA Nanostructure Dynamics: Shape, Self‐Complementarity and Spatial Flexibility
    ChemBioChem (IF 2.774) Pub Date : 2018-07-14
    Simon Chi-Chin Shiu; Andrew Brian Kinghorn; Yusuke Sakai; Yee-Wai Cheung; Jonathan Gardiner Heddle; Julian Alexander Tanner

    DNA aptamers are ideal tools to enable modular control of DNA nanostructure dynamics. For molecular recognition they have a particular advantage compared to antibodies, namely they can be integrated into DNA nanostructures in a bespoke manner by base‐pairing or nucleotide extension without any complex bioconjugation strategy. Such simplicity will be critical when considering advanced therapeutic and diagnostic applications of DNA nanostructures. However, optimizing DNA aptamers for functional control of DNA nanostructure dynamics can be challenging. Herein we present three considerations ‐ shape, self‐complementarity and spatial flexibility ‐ that should be paramount when optimizing aptamer functionality. These lessons, learnt from the growing number of aptamer‐nanostructure reports thus far, will be helpful for future studies where aptamers are used to control nucleic acid nanostructure dynamics.

    更新日期:2018-07-15
  • Quinoline–Glycomimetic Conjugates Reducing Lipogenesis and Lipid Accumulation in Hepatocytes
    ChemBioChem (IF 2.774) Pub Date : 2018-06-13
    Subhadeep Palit; Sanghamitra Mukherjee; Sougata Niyogi; Anindyajit Banerjee; Dipendu Patra; Amit Chakraborty; Saikat Chakrabarti; Partha Chakrabarti; Sanjay Dutta
    更新日期:2018-07-14
  • A Designed Enzyme Promotes Selective Post‐translational Acylation
    ChemBioChem (IF 2.774) Pub Date : 2018-07-10
    Pallavi M. Gosavi; Megha Jayachandran; Joel J. L. Rempillo; Oleksii Zozulia; Olga V. Makhlynets; Ivan V. Korendovych
    更新日期:2018-07-12
  • An Aminocaprolactam Racemase from Ochrobactrum anthropi with Promiscuous Amino Acid Ester Racemase Activity
    ChemBioChem (IF 2.774) Pub Date : 2018-06-13
    Amina Frese; Sarah V. Barrass; Peter W. Sutton; Joe P. Adams; Gideon Grogan
    更新日期:2018-07-12
  • Extending the Applicability of Exact Nuclear Overhauser Enhancements to Large Proteins and RNA
    ChemBioChem (IF 2.774) Pub Date : 2018-06-08
    Parker J. Nichols; Alexandra Born; Morkos A. Henen; Dean Strotz; Chi N. Celestine; Peter Güntert; Beat Vögeli
    更新日期:2018-07-12
  • Bioorthogonally Applicable Fluorescence Deactivation Strategy for Receptor Kinetics Study and Theranostic Pretargeting Approaches
    ChemBioChem (IF 2.774) Pub Date : 2018-06-04
    Steffen van der Wal; Clarize M. de Korne; Laurens G. L. Sand; Danny M. van Willigen; Pancras C. W. Hogendoorn; Karoly Szuhai; Fijs W. B. van Leeuwen; Tessa Buckle
    更新日期:2018-07-12
  • The Photosensitizing Clinical Agent Verteporfin is an Inhibitor of SPAK and OSR1 Kinases
    ChemBioChem (IF 2.774) Pub Date : 2018-07-12
    Mubarak A AlAmri; Hachemi Kadri; Mark Jeeves; Luke J Alderwick; Youcef Mehellou

    SPAK and OSR1 are two serine/threonine protein kinases that play important key roles in regulating ion homeostasis. Various SPAK and OSR1 mouse models exhibited reduced blood pressure. Herein, we report the discovery of Verteporfin, a photosensitizing agent used in photodynamic therapy, as a potent inhibitor of SPAK and OSR1 kinases. We show that Verteporfin binds the kinase domains of SPAK and OSR1 and inhibit their catalytic activity in an ATP‐independent manner. In cells, Verteporfin was able to suppress the phosphorylation of the ion co‐transporter NKCC1, a downstream physiological substrate of SPAK and OSR1 kinases. Kinase panel screening indicated that Verteporfin inhibited a further eight protein kinases more potently than SPAK and OSR1. Although Verteporfin has largely been studied as a modifier of the Hippo signaling pathway, this work indicates that the WNK‐SPAK/OSR1 signaling cascade is also a target of this clinical agent. This finding could explain the fluctuation in blood pressure noted in patients and animals treated with this drug.

    更新日期:2018-07-12
  • An In Vitro Selected DNAzyme Mutant Highly Specific for Na+ in Slightly Acidic Conditions
    ChemBioChem (IF 2.774) Pub Date : 2018-07-10
    Lingzi Ma; Juewen Liu

    Sodium is one of the most ubiquitous metal ions in biology, while its DNA‐based probes were reported only recently. A Na+‐specific RNA‐cleaving DNAzyme named NaA43 is active with Na+ alone. In this work, we used Co(NH3)63+ as the intended metal cofactor for in vitro selection, but obtained a mutant of the NaA43 DNAzyme. The mutant was named NaH1, which differs from NaA43 by only two nucleotides. NaA43 has an optimal of pH 7.0, while the optimal pH for NaH1 is 6.0. This difference might be due to our selection being performed at pH 6.0. Compared to other competing monovalent ions, NaH1 also displays an excellent selectivity for sodium and a fast catalytic rate of 0.11 ± 0.01 min‐1 with 50 mM Na+. At low Na+ concentrations, the selected DNAzyme exhibited a higher cleavage rate than NaA43 and thus a tighter apparent Kd of 12.0 ± 1.6 mM Na+. Furthermore, the NaH1 DNAzyme was engineered into a fluorescent Na+ biosensor by labeling a fluorophore/quencher pair that the sensor had a detection limit of 223 µM Na+. Preliminary work on detection Na+ in serum was demonstrated as well. This study provides a useful mutant that works in a slightly acidic environment, which might be useful for sensing Na+ in acidic in vivo environment.

    更新日期:2018-07-12
  • Mechanism of BRAF Activation Through Biochemical Characterization of the Recombinant Full‐Length Protein
    ChemBioChem (IF 2.774) Pub Date : 2018-07-10
    Nicholas Cope; Christine Candelora; Kenneth Wong; Sujeet Kumar; Haihan Nan; Michael Grasso; Borna Novak; Yana Li; Ronen Marmorstein; Zhihong Wang

    BRAF kinase plays an important role in mitogen activated protein kinase (MAPK) signaling and harbors activating mutations in about half of melanomas and in a smaller percentage in many other cancers. Despite its importance, few in vitro studies have been performed to characterize the biochemical properties of full‐length BRAF. Here we describe a strategy to generate an active, intact form of BRAF protein suitable for in vitro enzyme kinetics. We show that purified intact BRAF protein auto‐phosphorylates its kinase activation loop and this can be enhanced by binding its MEK protein substrate through an allosteric mechanism. Our studies provide in vitro evidence that BRAF selectively binds to active RAS and that the BRAF/CRAF heterodimer is the most active form relative to their respective homodimers. Full‐length BRAF analysis with small molecule BRAF inhibitors shows that two drugs, dabrafenib and vemurafenib, can modestly enhance BRAF's kinase activity at low concentration. Taken together, our characterization of intact BRAF contributes to a framework for understanding its role in cell signaling.

    更新日期:2018-07-12
  • Discovery of Antimicrobial Lipodepsipeptides Produced by a Serratia sp. within Mosquito Microbiomes
    ChemBioChem (IF 2.774) Pub Date : 2018-07-09
    Jack G. Ganley; Gavin Carr; Thomas R. Ioerger; James C. Sacchettini; Jon Clardy; Emily R. Derbyshire
    更新日期:2018-07-09
  • A Defined and Flexible Pocket Explains Aryl Substrate Promiscuity of the Cahuitamycin Starter Unit–Activating Enzyme CahJ
    ChemBioChem (IF 2.774) Pub Date : 2018-07-09
    Ashootosh Tripathi; Sung Ryeol Park; Andrew P. Sikkema; Hyo Je Cho; Jianfeng Wu; Brian Lee; Chuanwu Xi; Janet L. Smith; David H. Sherman
    更新日期:2018-07-09
  • 更新日期:2018-07-09
  • Flexible assembly of cascade enzyme on DNA triangle prism nanostructure for controlled biomimetic generation of nitric oxide
    ChemBioChem (IF 2.774) Pub Date : 2018-07-09
    Li Zhou; Yu Liu; Hui Shi; Xiaohai Yang; Jin Huang; Songyang Liu; Qiaoshu Chen; Jianbo Liu; Kemin Wang

    Abstract: Spatial organization of multienzyme at specific positions for controlled reaction cascade has attracted wide attentions. Here, we report the construction of a biomimetic enzyme cascade organizing on DNA triangle prism (TP) nanostructures, which enabled efficient catalytical production of nitric oxide (NO) on a single microbead. Two enzymes, glucose oxidase (GOx) and horseradish peroxidase (HRP), were assembled at adjacent locations on DNA TP nanostructure by using DNA‐binding protein adaptors with small interenzyme distances. In the cascade, enzyme GOx converts glucose into gluconic acid in the presence of oxygen. The produced intermediate H2O2 is rapidly transported to the enzyme HRP, which oxides hydroxyurea into NO and other nitroxyl species. The pH near the surface of the negatively charged DNA nanostructures is believed to be lower than that in the bulk solution, creating a more optimal pH environment for the anchored enzymes, resulted in a higher yields of the product NO molecules. Furthermore, the multienzyme system was immobilized on a microbead mediated by DNA adaptor, which enabled efficient catalytical generation of the gas molecules in the microreactor. Therefore, this work provides an alternative route for the biomimetic generation of NO through enzyme cascades. In particular, the dynamic binding capability of the DNA sequence enabled the reversible position of the protein enzyme and DNA nanostructure, which allowed to modulate the cascade catalysis.

    更新日期:2018-07-09
  • Antibody‐Bactericidal Macrocyclic Peptide Conjugates to Target Gram Negative Bacteria
    ChemBioChem (IF 2.774) Pub Date : 2018-07-08
    Faycal Touti; Guillaume Lautrette; Ken Johnson; Jim Delaney; Andrew Wollacott; Hamid Tissire; Karthik Viswanathan; Zachary Shriver; Surin Mong; Alexander Mijalis; Obadiah Plante; Bradley Pentelute

    To combat antimicrobial infections, new active molecules are needed. Antimicrobial peptides, ever abundant in nature, are a fertile starting point to develop new antimicrobial agents but suffer from low stability, low specificity and off‐target toxicity. These drawbacks have limited their development. To overcome some of these limitations, we developed antibody‐bactericidal macrocyclic peptide conjugates (ABCs) where the antibody directs the bioactive macrocyclic peptide to the targeted Gram‐negative bacteria. We used cysteine SNAr chemistry to synthesize and systematically study a library of large (>30 mer) macrocyclic antimicrobial peptides (mAMPs) to discover variants with extended proteolytic stability in human serum and low hemolytic activity while maintaining bioactivity. We then conjugated, using sortase A, these bioactive variants onto an E. coli targeted monoclonal antibody. We found these ABCs had minimized hemolytic activity and are able to kill E. coli at nanomolar concentrations. Our findings suggest macrocyclic peptides when fused to antibodies may facilitate the discovery of new agents to treat bacterial infections.

    更新日期:2018-07-09
  • Ultrastructural Imaging of Salmonella–Host Interactions Using Super‐resolution Correlative Light‐Electron Microscopy of Bioorthogonal Pathogens
    ChemBioChem (IF 2.774) Pub Date : 2018-06-05
    Daphne M. van Elsland; Sílvia Pujals; Thomas Bakkum; Erik Bos; Nikolaos Oikonomeas‐Koppasis; Ilana Berlin; Jacques Neefjes; Annemarie H. Meijer; Abraham J. Koster; Lorenzo Albertazzi; Sander I. van Kasteren
    更新日期:2018-07-08
  • Enzymatically Ligated DNA–Surfactants: Unmasking Hydrophobically Modified DNA for Intracellular Gene Regulation
    ChemBioChem (IF 2.774) Pub Date : 2018-06-03
    Alyssa K. Hartmann; Dominic F. Cairns‐Gibson; Joshua J. Santiana; Mark Q. Tolentino; Halle M. Barber; Jessica L. Rouge
    更新日期:2018-07-08
  • Diversity‐Oriented Synthesis of Diol‐Based Peptidomimetics as Potential HIV Protease Inhibitors and Antitumor Agents
    ChemBioChem (IF 2.774) Pub Date : 2018-06-02
    Paresh M. Vadhadiya; Marc‐Alexandre Jean; Chahrazed Bouzriba; Thomas Tremblay; Patrick Lagüe; Sébastien Fortin; John Boukouvalas; Denis Giguère
    更新日期:2018-07-08
  • Unravelling the Specificity of Laminaribiose Phosphorylase from Paenibacillus sp. YM‐1 towards Donor Substrates Glucose/Mannose 1‐Phosphate by Using X‐ray Crystallography and Saturation Transfer Difference NMR Spectroscopy
    ChemBioChem (IF 2.774) Pub Date : 2018-06-01
    Sakonwan Kuhaudomlarp; Samuel Walpole; Clare E. M. Stevenson; Sergey A. Nepogodiev; David M. Lawson; Jesus Angulo; Robert A. Field
    更新日期:2018-07-08
  • Artificial Light Regulation of an Allosteric Bienzyme Complex by a Photosensitive Ligand
    ChemBioChem (IF 2.774) Pub Date : 2018-05-29
    Andrea C. Kneuttinger; Martin Winter; Nadja A. Simeth; Kristina Heyn; Rainer Merkl; Burkhard König; Reinhard Sterner
    更新日期:2018-07-08
  • Protein Engineering of the Progesterone Hydroxylating P450‐Monooxygenase CYP17A1 Alters its Regioselectivity
    ChemBioChem (IF 2.774) Pub Date : 2018-07-07
    Lisa Morlock; Sascha Grobe; Kathleen Balke; Stephan Mauersberger; Dominique Böttcher; Uwe Bornscheuer

    The CYP171 enzyme is known to catalyse a key step in the steroidogenesis of mammals. The substrates progesterone and pregnenolone are first hydroxylated at the C17 position, followed by the cleavage of the C17‐C20 bond, yielding important precursors for glucosteroids and androgens. In this study, we focused on the reaction of the bovine CYP17A1 enzyme with progesterone as substrate. On the basis of a created homology model, active‐site residues were identified and, systematically mutated to alanine. In whole‐cell biotransformations, the importance of the residues N202, R239, G297, and E305 for substrate conversion was confirmed. Additionally, the mutation of the residues L206, V366, and V483 enhanced the formation of the side‐product 16α‐hydroxyprogesterone up to 40% of total product formation. Furthermore, residue L105 was found not being involved in this side‐activity, contradicting a former study with the human enzyme.

    更新日期:2018-07-08
  • Azide‐ and alkyne‐bearing metabolic chemical reporters of glycosylation show structure‐dependent feedback inhibition of the hexosamine biosynthetic pathway.
    ChemBioChem (IF 2.774) Pub Date : 2018-07-06
    Lisa A Walter; Anna R Batt; Narek Darabedian; Balyn W Zaro; Matthew Robert Pratt

    Metabolic chemical reporters (MCRs) of protein glycosylation are analogs of natural monosaccharides that bear reactive groups, like azides and alkynes. Upon treatment of living cells and organisms, these small molecules are biosynthetically transformed into nucleotide donor‐sugars and then utilized by glycosyltransferases for the modification of proteins. Subsequent installation of tags using bioorthogonal chemistries can then enable the visualization and enrichment of these glycoproteins. While this two‐step procedure is powerful, the utilization of MCRs has the potential to change the endogenous production of the natural repertoire of donor sugars. A major route for the generation of these glycosyltransferase substrates is the hexosamine biosynthetic pathway (HBP) that results in uridine diphosphate N‐acetylglucosamine (UDP‐GlcNAc). Interestingly, the rate determining enzyme of the HBP, glutamine fructose‐6‐phosphate amidotransferase (GFAT), is feedback inhibited by UDP‐GlcNAc. This raises the possibility that the build‐up of UDP‐MCRs will block the biosynthesis of UDP‐GlcNAc, resulting in off target effects. Here, we directly test this possibility with recombinant human GFAT and a small panel of synthetic UDP‐MCRs. We find that MCRs with larger substitutions at the N‐acetyl position do not inhibit GFAT, while those with modifications of the 2‐ or 6‐hydroxyl maintain this ability. These results further illuminate the considerations that should be applied to the use of MCRs.

    更新日期:2018-07-08
  • DNA‐binding properties of new fluorescent AzaHx‐amides: Methoxy‐pyridyl‐aza‐benzimidazole‐pyrrole‐imidazole/pyrrole
    ChemBioChem (IF 2.774) Pub Date : 2018-07-04
    Beibei Liu; Luke Pett; Konstantinos Kiakos; Pravin C. Patil; Vijay Satam; John A. Hartley; Moses Lee; W. David Wilson

    DNA minor groove binding polyamides have been extensively developed to control abnormal gene expression. Establishment of the novel, inherently fluorescent Hx‐amides has provided an alternative path for studying DNA binding in cells by direct observation of cell localization. Because of the 2:1 antiparallel stacking homodimer binding mode of these molecules to DNA, modification of Hx‐amides to AzaHx‐amides has successfully extended the DNA recognition repertoire, from central CG (recognized by Hx‐I) to central GC (recognized by AzaHx‐P) recognition. To potentially target two consecutive GG bases, new modifications from AzaHx moiety to 3‐Pyr‐AzaHx and 2‐Pyr‐AzaHx moieties were developed. The newly designed molecules are also small‐sized, fluorescent amides with Pyr‐AzaHx connected to two conventional five‐membered heterocycles. Complementary biophysical methods were carried out to investigate DNA binding properties of these molecules. The results showed that neither 3‐Pyr‐AzaHx nor 2‐Pyr‐AzaHx was able to mimic I‐I to specifically target GG dinucleotides. Rather, 3‐Pyr‐AzaHx functions like AzaHx or f‐I or P‐I as a antiparallel stacked dimer. 3‐Pyr‐AzaHx‐PI (2) binds 5'‐ACGCGT'‐3' with improved binding affinity and high sequence specificity when compared to its parent molecule AzaHx‐PI (1). However, 2‐Pyr‐AzaHx is detrimental to DNA binding because of an unfavorable steric clash upon stacking in the minor groove.

    更新日期:2018-07-08
  • Biosynthetic Gene Cluster of a D‐Tryptophan‐Containing Lasso Peptide, MS‐271
    ChemBioChem (IF 2.774) Pub Date : 2018-07-04
    Zhi Feng; Yasushi Ogasawara; Satoshi Nomura; Tohru Dairi

    MS‐271, produced by Streptomyces sp. M‐271, is a lasso peptide natural product comprising 21 amino acid residues with a D‐tryptophan (D‐Trp) at its C‐terminus. Because lasso peptides are ribosomal peptides, the biosynthesis of MS‐271, especially the mechanism of D‐Trp introduction, is of great interest. The MS‐271 biosynthetic gene cluster was identified by draft genome sequencing of the MS‐271 producer and it was revealed that the precursor peptide contains all 21 amino acid residues including the C‐terminal tryptophan. This suggested that the D‐Trp residue is introduced via epimerization. Modification enzyme genes such as a macrolactam synthetase (MslC), precursor peptide recognition element (MslB1), cysteine protease (MslB2), disulfide oxidoreductases (MslE, MslF), and a function‐unknown protein (MslH) were found in the flanking region of the precursor peptide gene. Although obvious epimerase genes were absent in the cluster, heterologous expression of the putative MS‐271 cluster in Streptomyces lividans showed that it contains all the necessary genes for MS‐271 production including a gene for a novel peptide epimerase. Furthermore, a gene deletion experiment indicated that MslB1, B2, C and H were indispensable for MS‐271 production and that some interactions of the biosynthetic enzymes were essential for the biosynthesis of MS‐271.

    更新日期:2018-07-08
  • A Glutathione‐Responsive Short Sequence of Metal–Organic Complex Array
    ChemBioChem (IF 2.774) Pub Date : 2018-05-28
    Purnandhu Bose; Toshiaki Takei; Xianglan Li; Takashi Minowa; Rajamani Rajmohan; Pothiappan Vairaprakash; Kentaro Tashiro
    更新日期:2018-07-04
  • DNA primer extension with cyclopropenylated 7‐deaza‐2'‐deoxyadenosine and efficient bioorthogonal labeling in vitro and in living cells
    ChemBioChem (IF 2.774) Pub Date : 2018-07-03
    Damian Damian Ploschik; Franziska Rönicke; Hanna Beike; Ralf Strasser; Hans-Achim Wagenknecht

    A dATP analog for DNA labeling was synthesized with the 1‐methylcylopropene (1MCP) group at the 7‐position of 7‐deaza‐2'‐deoxyadenosine, and applied for primer extension experiments. The real‐time kinetic data reveals that this 1MCP‐modified dATP analog is much faster incorporated into DNA than the similarly 1MCP‐modified dUTP analog. The postsynthetic fluorescent labeling of these oligonucleotides works efficient according to PAGE analysis, and can be applied for immobilization of a functional antibody on a surface. Site‐specific labeling at two different positions in DNA was achieved and the bioorthogonality of the postsynthetic fluorescent labeling was demonstrated in living HeLa cells.

    更新日期:2018-07-04
  • Characterisation of the Carboxypeptidase G2 Catalytic Site and Design of New Inhibitors for Cancer Therapy
    ChemBioChem (IF 2.774) Pub Date : 2018-07-03
    Dhadchayini Jeyaharan; Carla Brackstone; James Schouten; Paul Davis; Ann M. Dixon

    The enzyme carboxypeptidase G2 (CPG2) is utilized in antibody directed enzyme pro‐drug therapy (ADEPT) to catalyze the formation of an active drug from an inert pro‐drug. Free CPG2 in the bloodstream must be inhibited before administration of the pro‐drug in order to avoid a systemic reaction in the patient. Although a few small‐molecule CPG2 inhibitors have been reported, none have been taken forward thus far. This lack of progress is due in part to a lack of structural understanding of the CPG2 active site as well as the absence of small molecules that can block the active site whilst targeting the complex for clearance. The work described here aimed to address both areas. We report the structural/functional impact of extensive point mutation across the putative CPG2 catalytic site and adjacent regions for the first time, revealing residues outside the catalytic region (K208A, S210A and T357A) crucial to enzyme activity. We also describe novel molecules that inhibit CPG2 whilst maintaining accessibility of galactosylated moieties aimed at targeting the enzyme for clearance. This work acts as a platform for future development of high‐affinity CPG2 inhibitors that occupy new chemical space and will advance the safe application of ADEPT in cancer treatment.

    更新日期:2018-07-04
  • Kinetically Stable Bicelle with Dilution Tolerance, Size Tunability, and Thermoresponsiveness for Drug Delivery Applications
    ChemBioChem (IF 2.774) Pub Date : 2018-07-03
    Noriyuko Uchida; Noriko Nishizawa Horimoto; Kuniyo Yamada; Takaaki Hikima; Yasuhiro Ishida

    Mixtures of a phospholipid (1,2‐dipalmitoyl‐sn‐glycero‐3‐phosphatidylcholine: DPPC) and a sodium cholate‐derived surfactant (SC‐C5) at room temperature formed phospholipid bilayer fragments that were edge‐stabilized by SC‐C5 so‐called bicelles. Because the bilayer melting point of DPPC (41 °C) is above room temperature and because SC‐C5 has an exceptionally low critical micelle concentration (<0.5 mM), the bicelles are kinetically frozen at room temperature. Consequently, they exist even when the mixture is diluted to a concentration of 0.04 wt%. In addition, the lateral size of the bicelles can be fine‐tuned by altering the molar ratio of DPPC to SC‐C5. On heating to ~37 °C, the bicelles transformed into micelles composed of DPPC and SC‐C5. By taking advantage of the dilution tolerance, size tunability, and thermoresponsiveness, we demonstrated in vitro drug delivery using the bicelles as carriers, which suggests their potential utility in transdermal drug delivery.

    更新日期:2018-07-04
  • Enzymatic fluorination of biotin and tetrazine conjugates for pre‐targeting approaches to PET imaging
    ChemBioChem (IF 2.774) Pub Date : 2018-07-02
    Phillip T Lowe; Sergio Dall'Angelo; Andrew Devine; Matteo Zanda; David O'Hagan

    The use of radiolabelled antibodies and antibody‐derived recombinant constructs have shown promise for both imaging and therapeutic use. In this context, the biotin‐avidin/streptavidin pairing along with the inverse electron demand Diels‐Alder reaction (IEDDA) have found application in pre‐targeting approaches for positron emission tomography (PET). This study reports the fluorinase enzyme mediated transhalogenation (ClDA substrates to FDA products) of two antibody pre‐targetting tools, a FDA‐PEG‐tetrazine and a [18F]FDA‐PEG‐biotin and each is assessed for their compatibility towards IEDDA ligation to trans‐cyclooctene or affinity to avidin. A protocol was developed to avoid radiolytically promoted oxidation of biotin during the synthesis of [18F]FDA‐PEG‐biotin. The study adds to the repertoire of bioconjugates for use in fluorinase catalysed radio‐synthesis for PET and shows that the fluorinase will accept a wide range of ClDA substrates tethered at C‐2 of the adenine ring with a pegylated cargo. The method is exceptional as the nucleophilic reaction with [18F]fluoride takes place in water at neutral pH and at ambient temperature.

    更新日期:2018-07-02
  • A possible path to prebiotic peptides involving silica and hydroxy acid‐mediated amide bond formation
    ChemBioChem (IF 2.774) Pub Date : 2018-06-29
    Aaron D McKee; Thomas Michael Orlando; Nicholas V Hud; Christopher Bennett; Andrew K Saydjari; Martin Solano

    Formation of alanine and glycine oligomers in films produced by drying aqueous mixtures of lactic acid and silica nanoparticles have been studied under plausible prebiotic conditions. The addition of silica results in alanine or glycine enrichment in the polymers. Oligomerization proceeds via ester‐mediated peptide bond formation in an acidic and evaporative environment at temperatures as low as 85 °C. For both amino acids, the dominant species produced in the presence of silica and lactic acid are rich in amide bonds and ester deficient. At higher temperatures, glycine and alanine oligomers contain only a single hydroxy acid residue conjugated to the N‐terminus. Similar product distributions occurs with silica particles pre‐reacted with lactic acid, suggesting a catalytic role of a functionalized surface. This work highlights the role minerals may have served in transitioning from oligomers with both ester and amide linkages (depsipeptides) to peptides in a prebiotic context.

    更新日期:2018-07-01
  • Coordination‐Assisted Bioorthogonal Chemistry: Orthogonal Tetrazine Ligation with Vinylboronic Acid and a Strained Alkene
    ChemBioChem (IF 2.774) Pub Date : 2018-05-28
    Selma Eising; Bo‐Tao Xin; Fleur Kleinpenning; Jurriaan J. A. Heming; Bogdan I. Florea; Herman S. Overkleeft; Kimberly M. Bonger
    更新日期:2018-06-30
  • Discovery of the Tiancilactone Antibiotics by Genome Mining of Atypical Bacterial Type II Diterpene Synthases
    ChemBioChem (IF 2.774) Pub Date : 2018-05-27
    Liao‐Bin Dong; Jeffrey D. Rudolf; Ming‐Rong Deng; Xiaohui Yan; Ben Shen
    更新日期:2018-06-30
  • Protein Regulation by Intrinsically Disordered Regions: A Role for Subdomains in the IDR of the HIV‐1 Rev Protein
    ChemBioChem (IF 2.774) Pub Date : 2018-05-23
    Ofrah Faust; Dana Grunhaus; Odelia Shimshon; Eylon Yavin; Assaf Friedler
    更新日期:2018-06-30
  • Incorporation of azide groups into DNA using membrane‐permeable nucleotide triesters in vivo
    ChemBioChem (IF 2.774) Pub Date : 2018-06-28
    Masayuki Tera; Stella Glasauer; Nathan William Luedtke

    Metabolic incorporation of bioorthogonal functional groups into cellular nucleic acids can be impeded by insufficient phosphorylation of nucleosides. Previous studies found that 5‐(azidomethyl)‐2'‐deoxyuridine (AmdU) was incorporated into the DNA of HeLa cells expressing a low‐fidelity thymidine kinase, but not by wild‐type HeLa cells. Here we report that membrane‐permeable phosphotriester derivatives of AmdU can exhibit enhanced incorporation into the DNA of wild‐type cells and animals. AmdU monophosphate derivatives carrying either 5'‐bispivaloyloxymethyl (POM), 5'‐bisacetoxybenzyl (AB), or "Protide" protective groups were used to mask the phosphate group of AmdU prior to its entry into cells. The POM derivative "POM‐AmdU" exhibited superior chemical stability, greater metabolic incorporation efficiency, and lower toxicity than "AB‐AmdU". Remarkably, the addition of POM‐AmdU to the water of zebrafish larvae enabled the biosynthesis of azide‐modified DNA throughout the body.

    更新日期:2018-06-30
  • Super‐Resolution Imaging of Amyloid Structures over Extended Times Using Transient Binding of Single Thioflavin T Molecules
    ChemBioChem (IF 2.774) Pub Date : 2018-06-28
    Kevin Spehar; Tianben Ding; Yuanzi Sun; Niraja Kedia; Jin Lu; George R Nahass; Matthew D Lew; Jan Bieschke

    Oligomeric amyloid structures are crucial therapeutic targets in Alzheimer's and other amyloid diseases. However, these oligomers are too small to be resolved by standard light microscopy. We have developed a simple and versatile tool to image amyloid structures using Thioflavin T without the need for covalent labeling or immunostaining. Dynamic binding of single dye molecules generates photon bursts that are used for fluorophore localization on a nanometer scale. Thus, photobleaching cannot degrade image quality, allowing for extended observation times. Super‐resolution Transient Amyloid Binding (TAB) microscopy promises to directly image native amyloid using standard probes and record amyloid dynamics over minutes to days. We imaged amyloid fibrils from multiple polypeptides, oligomeric, and fibrillar structures formed during different stages of amyloid‐β aggregation, as well as the structural remodeling of amyloid‐β fibrils by the compound epi‐gallocatechin gallate (EGCG).

    更新日期:2018-06-30
  • Protein Crystallography and Site‐Direct Mutagenesis Analysis of the Poly(ethylene terephthalate) Hydrolase PETase from Ideonella sakaiensis
    ChemBioChem (IF 2.774) Pub Date : 2018-06-27
    Bing Liu; Lihui He; Liping Wang; Tao Li; Changcheng Li; Huayi Liu; Yunzi Luo; Rui Bao
    更新日期:2018-06-28
  • Auroramycin: A Potent Antibiotic from Streptomyces roseosporus by CRISPR‐Cas9 Activation
    ChemBioChem (IF 2.774) Pub Date : 2018-05-25
    Yee Hwee Lim; Fong Tian Wong; Wan Lin Yeo; Kuan Chieh Ching; Yi Wee Lim; Elena Heng; Shuwen Chen; De‐Juin Tsai; Tsai‐Ling Lauderdale; Kak‐Shan Shia; Ying Swan Ho; Shawn Hoon; Ee Lui Ang; Mingzi M. Zhang; Huimin Zhao
    更新日期:2018-06-28
  • 更新日期:2018-06-28
  • Synthesis, Characterisation and Reactivity of 3‐Mercaptopyruvic Acid
    ChemBioChem (IF 2.774) Pub Date : 2018-05-20
    Erwan Galardon; Jean‐Christophe Lec
    更新日期:2018-06-28
  • A split‐intein‐based method for the efficient production of circularized nanodiscs for structural studies of membrane proteins
    ChemBioChem (IF 2.774) Pub Date : 2018-06-27
    Miehling Jonas; David Goricanec; Franz Hagn

    Phospholipid nanodiscs are a native‐like membrane mimetic that is suitable for structural studies of membrane proteins. Even though nanodiscs of different sizes exist for various structural applications, their thermal and long‐term stability can vary considerably. Covalently circularized nanodiscs are a perfect tool to overcome these limitations. Existing methods for the production of circularized nanodiscs can be time consuming and technically demanding. Therefore, we here present an easy in vivo approach, where circularized MSP proteins can be directly obtained from E. coli culture. We use Npu DnaE split‐intein fusions with MSPs of various lengths and are able to consistently obtain circularized nanodiscs in high yields. Using this approach, we were able to produce a large variety of circularized nanodiscs ranging from 7 to 26 nm in diameter that are suitable for NMR as well as EM applications. These nanodiscs are superior to corresponding linear versions in terms of stability and size homogeneity, affecting the quality of NMR data as well as EM experiments. Due to their long‐term stability and homogeneity the presented small circular nanodiscs are well‐suited for high‐resolution NMR studies as demonstrated with two membrane proteins of 17 or 32 kDa in size. The presented method will provide easy access to circularized nanodiscs for structural studies of membrane proteins as well as for applications where a defined and stable nanodisc size is required.

    更新日期:2018-06-28
  • Highly Efficient Cell Penetrable Probes of Protein Arginine Deiminases (PADs) for Functional Proteomics
    ChemBioChem (IF 2.774) Pub Date : 2018-06-27
    Sanjai Kumar

    Protein Arginine Deiminases (PADs) have recently emerged at the forefront of drug discovery programs in several human disorders. Despite this, precise understanding of their functional roles in human biology remain poorly defined. This report highlights a recent development of next generation activity‐based PAD probes that are highly efficient, cell active, and metabolically stable. This discovery will rapidly expedite functional assignments of PAD biology in both normal and diseased cells, thereby leading the pathway for development of PAD‐targeted therapeutics in the near future.

    更新日期:2018-06-28
  • Characterization and reshaping of a large and hydrophobic nucleophile pocket in lipases/acyltransferases
    ChemBioChem (IF 2.774) Pub Date : 2018-06-27
    Anne-Helene Jan Deniau; Maeva Subileau; Eric Dubreucq

    Lipases/acyltransferases, such as CpLIP2 from Candida parapsilosis and CduLAc from Candida dubliniensis catalyze preferentially acyltransfer over hydrolysis when a suitable nucleophile is present, even in medium with a high thermodynamic activity of water (aW). They are related to CAL‐A from Moesziomyces antarcticus, which in comparison displays a lower acyltransfer ability. The 3D structures of wild‐types and mutants of CAL‐A, CpLIP2 and CduLAc revealed differences in size and hydrophobicity of a large pocket located under the catalytic triad. The kinetic behavior of site‐directed mutants confirmed the role of this pocket in the competition between methanol and water as the nucleophile acceptor for the deacylation step. The mutations provided a better understanding of key structural determinants for the variable levels of acyltransferase ability observed and supported the existence of a complex network of nucleophile interactions within the enzymes. The shape and size of the possible nucleophile pocket identified also suggested that multiple binding sites could exist, supporting the hypothesis of non‐overlapping leaving and accepting nucleophiles binding sites.

    更新日期:2018-06-28
  • Biosynthetic and Functional Color–Scent Associations in Flowers of Papaver nudicaule and Their Impact on Pollinators
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    Jaime Martínez‐Harms; Anne‐Christin Warskulat; Bettina Dudek; Grit Kunert; Sybille Lorenz; Bill S. Hansson; Bernd Schneider
    更新日期:2018-06-27
  • Site‐Specific Incorporation of Chemical Fluorescence on Live Enterovirus‐71 Virion by Using an Organometallic Palladium Reagent To Monitor Virus Entry
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    Yaxin Wang; Jingwei Liu; Lin Cao; Wenjing Wang; Yuna Sun; Zheng Yin; Zhiyong Lou
    更新日期:2018-06-27
  • Preparation of Cyclic Prodiginines by Mutasynthesis in Pseudomonas putida KT2440
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    Andreas Sebastian Klein; Hannah Ursula Clara Brass; David Paul Klebl; Thomas Classen; Anita Loeschcke; Thomas Drepper; Sonja Sievers; Karl‐Erich Jaeger; Jörg Pietruszka
    更新日期:2018-06-27
  • Osmium Tag for Post‐transcriptionally Modified RNA
    ChemBioChem (IF 2.774) Pub Date : 2018-05-25
    Turja Kanti Debnath; Akimitsu Okamoto
    更新日期:2018-06-27
  • A Redox‐Based Superoxide Generation System Using Quinone/Quinone Reductase
    ChemBioChem (IF 2.774) Pub Date : 2018-05-23
    Shailesh Kumar Singh; Syed Masood Husain
    更新日期:2018-06-27
  • Synthesis and Characterization of a New Bifunctionalized, Fluorescent, and Amphiphilic Molecule for Recruiting SH‐Containing Molecules to Membranes
    ChemBioChem (IF 2.774) Pub Date : 2018-05-22
    Monique Mertens; Malte Hilsch; Ivan Haralampiev; Rudolf Volkmer; Pablo Wessig; Peter Müller
    更新日期:2018-06-27
  • The Photophysics of Polythiophene Nanoparticles for Biological Applications
    ChemBioChem (IF 2.774) Pub Date : 2018-05-01
    Ilaria Bargigia; Elena Zucchetti; Ajay Ram Srimath Kandada; Miguel Moreira; Caterina Bossio; Walter P. D. Wong; Paulo Barbeitas Miranda; Paolo Decuzzi; Cesare Soci; Cosimo D'Andrea; Guglielmo Lanzani
    更新日期:2018-06-27
  • Co‐immobilized whole‐cells with ω‐transaminase and ketoreductase activity for continuous‐flow cascade reaction
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    László Nagy-Győr; Emese Abaházi; Viktória Bódai; Péter Sátorhelyi; Balázs Erdélyi; Diána Balogh-Weiser; Csaba Paizs; Gábor Hornyánszky; Laszlo Poppe

    An improved sol‐gel process using hollow silica microspheres as supporting additive was applied for co‐immobilization of whole‐cells of E. coli with Chromobacterium violaceum ω‐transaminase activity and Lodderomyces elongisporus with ketoreductase activity. The co‐immobilized cells with two different biocatalytic activity could perform a reaction cascade converting racemic 4‐phenylbutan‐2‐amine or heptan‐2‐amine to a nearly equimolar mixture of enantiomerically pure (R)‐amine and the corresponding (S)‐alcohol even in continuous‐flow mode. The novel co‐immobilized whole‐cell system proved to be an easy‐to‐store and durable biocatalyst.

    更新日期:2018-06-27
  • A One‐pot ''Triple‐C'' Multicyclization Methodology for the Synthesis of Highly Constrained Isomerically Pure Tetracyclic Peptides
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    Gaston Richelle; Marcel Schmidt; Hans Ippel; Tilman Hackeng; Jan van Maarseveen; Timo Nuijens; Peter Timmerman

    Here we report a broadly applicable one‐pot methodology for the facile transformation of linear peptides into tetracyclic peptides via a CEPS/CLIPS/CuAAC ("triple‐C") locking methodology. Linear peptides with varying lengths (≥ 14 amino acids), comprising two cysteines and two azidohomoalanines (Aha), were efficiently cyclized head‐to‐tail using the peptiligase variant omniligase‐1 (CEPS). Subsequent ligation‐cyclization using tetravalent (T41/2) scaffolds containing two bromomethyl groups and two alkyne functionalities yielded isomerically pure tetracyclic peptides. Sixteen different functional tetracycles, derived from bicyclic inhibitors against urokinase plasminogen activator (uPA) and coagulation factor XIIa (FXIIa), were successfully synthesized and their bioactivities evaluated. Two of these (FF‐T41/2) exhibited increased inhibitory activity against FXIIa as compared to a bicyclic control peptide. The corresponding hetero‐bifunctional variants (UF/FU‐T41/2), bearing a single copy of each inhibitory sequence, exhibited micromolar activities against both uPA and FXIIa, thus illustrating the potential of the ''bifunctional tetracycle peptide'' inhibitor concept.

    更新日期:2018-06-27
  • High‐Throughput Small‐Molecule Enantiopurity Measurement Using Flow Cytometry
    ChemBioChem (IF 2.774) Pub Date : 2018-06-26
    Zhesen Tan; Jennifer Heemstra

    Fluorescence‐activated cell sorting (FACS) offers a powerful approach to high‐throughput library screening in directed evolution experiments. However, FACS is rarely utilized in the evolution of stereoselective enzymes, due to the difficulty of designing fluorescence‐based assays for measuring enantiopurity. Here, we describe a novel FACS‐based enantiopurity analysis approach that overcomes these limitations by utilizing enantiomeric DNA biosensors labeled with orthogonal fluorophores. By co‐encapsulating the biosensors with a mixture of target enantiomers in microfluidic droplets, we demonstrate the use of FACS to differentiate between droplets having varying levels of target enantiopurity. We envision the utility of this method for high‐throughput screening of enantiopurity in the directed evolution of stereoselective enzymes, facilitating the discovery of new asymmetric biocatalysts for the synthesis of pharmaceuticals and other high‐value chemicals.

    更新日期:2018-06-27
  • The Red‐/Green‐switching GAF3 of cyanobacteriochrome Slr1393 from Synechocystis sp. PCC6803 regulates the activity of an adenylyl cyclase
    ChemBioChem (IF 2.774) Pub Date : 2018-06-25
    Ping-Ping Hu; Rui Guo; Ming Zhou; Wolfgang Gärtner; Kai-Hong Zhao

    Cyanobacteriochromes (CBCRs) are photoreceptors in cyanobacteria that present a bilin chromophore‐binding GAF domain as a photochromic element to control the activity of a downstream enzyme or regulator. CBCR Slr1393 from Synechocystis PCC 6803 carries three GAF domains, but only the third one binds covalently phycocyanobilin. Slr1393 shows photochromicity between a red‐ and a green absorbing state and regulates a C‐terminally located histidine kinase. In this work, we fused this third GAF domain to an adenylyl cyclase (AC) from Microcoleus chthonoplastes PCC7420 that in its genuine form is under blue light control from a LOV domain. A series of RGS‐AC variants were constructed with various length of linkers between RGS and AC. Assays in vitro and in living Escherichia coli cells (AC‐deletion mutant) demonstrate that the activity of AC is light‐regulated, namely the red light converted form of RGS∆14‐∆4AC (in vitro) was about three‐fold more active than the green light converted form. Expression of the fusion protein RGS∆14‐∆4AC in vivo again showed highest light regulation with an at least three fold amplification of the AC function. In some experiments yielded even ten‐fold higher activity indicating that the protein when expressed under in vivo conditions is part of the E. coli physiological conditions and thereby subject to more complex and variable regulation through other E. coli‐inherent factors.

    更新日期:2018-06-27
  • Covalent chemical co‐chaperones of the p300/CBP GACKIX domain
    ChemBioChem (IF 2.774) Pub Date : 2018-06-25
    Jean M Lodge; Chinmay Y Majmudar; James Clayton; Anna Kathryn Mapp

    The GACKIX activator‐binding domain has been a compelling target for small molecule probe discovery because of the central role activator‐GACKIX complexes play in diseases ranging from leukemia to memory disorders. Additionally, GACKIX is an ideal model to dissect the context‐dependent function of activator‐coactivator complexes. However, the dynamic and transient PPIs formed by GACKIX are difficult targets for small molecules. An additional complication is that activator‐binding motifs such as GACKIX are found in multiple coactivators, making specificity difficult to attain. In this study, we demonstrate that the strategy of Tethering can be used to rapidly discover highly specific covalent modulators of the dynamic PPIs between activators and coactivators. These serve as both orthosteric and allosteric modulators, enabling the tunable assembly or disassembly of the activator‐coactivator complexes formed between the KIX domain and its cognate activator binding partners MLL and CREB. The molecules maintain their function and selectivity even in human cell lysates and in bacterial cells and thus will ultimately be useful probes for cellular studies.

    更新日期:2018-06-27
  • A bioluminescent Ca2+ indicator based on a topological variant of GCaMP6s
    ChemBioChem (IF 2.774) Pub Date : 2018-06-22
    Yong Qian; Vladimir Rancic; Jiahui Wu; Klaus Ballanyi; Robert E Campbell

    Fluorescent genetically‐encoded calcium ion (Ca2+) indicators (GECIs) enable monitoring of Ca2+ dynamics in a diverse array of cell types and tissues. One drawback of green fluorescent GECIs, such as the widely used GCaMP6 variant, is that the blue wavelengths of light used to excite the GECI also activate optogenetic actuators such as channelrhodopsins. Accordingly, it is particularly challenging to simultaneously use both optogenetic actuators and GECIs to both control and image cell signaling. Bioluminescence is an alternative imaging modality that circumvents this problem by avoiding the use of illumination for fluorescence excitation. Here, we report the development of a bioluminescent GECI, designated LUCI‐GECO1, based on efficient bioluminescent resonance energy transfer (BRET) between the NanoLuc luciferase and a topological variant of GCaMP6s. LUCI‐GECO1 is a sensitive ratiometric GECI that retains the highly optimized properties of GCaMP6s, as we demonstrate by imaging of chemically and optogenetically‐induced Ca2+ concentration changes in cultured cells and neurons.

    更新日期:2018-06-25
  • Proline Fingerprint in Intrinsically Disordered Proteins
    ChemBioChem (IF 2.774) Pub Date : 2018-05-23
    Maria Grazia Murrali; Alessandro Piai; Wolfgang Bermel; Isabella C. Felli; Roberta Pierattelli
    更新日期:2018-06-22
  • Cyclic Peptides for Efficient Detection of Collagen
    ChemBioChem (IF 2.774) Pub Date : 2018-05-14
    Koh K. Takita; Kazunori K. Fujii; Tetsuya Kadonosono; Ryo Masuda; Takaki Koide
    更新日期:2018-06-22
  • A Defined and Flexible Pocket Explains Aryl Substrate Promiscuity of the Cahuitamycin Starter Unit–Activating Enzyme CahJ
    ChemBioChem (IF 2.774) Pub Date : 2018-05-09
    Ashootosh Tripathi; Sung Ryeol Park; Andrew P. Sikkema; Hyo Je Cho; Jianfeng Wu; Brian Lee; Chuanwu Xi; Janet L. Smith; David H. Sherman
    更新日期:2018-06-22
  • The Alkylquinolone Repertoire of Pseudomonas aeruginosa is Linked to Structural Flexibility of the FabH‐like 2‐Heptyl‐3‐hydroxy‐4(1H)‐quinolone (PQS) Biosynthesis Enzyme PqsBC
    ChemBioChem (IF 2.774) Pub Date : 2018-05-03
    Florian Witzgall; Tobias Depke; Michael Hoffmann; Martin Empting; Mark Brönstrup; Rolf Müller; Wulf Blankenfeldt
    更新日期:2018-06-22
Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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