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  • Covalent Modification of Biomolecules through Maleimide-Based Labeling Strategies
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-16
    Kévin Renault, Jean Wilfried Fredy, Pierre-Yves Renard, Cyrille Sabot
    更新日期:2018-07-16
  • 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
  • Immune Profiling of Polysaccharide Submicron Vesicles
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-16
    Chiara Cristina TOMA, Alessandra Aloisi, Valentina Bordoni, Riccardo Di Corato, Martina Rauner, Gianaurelio Cuniberti, Lucia Gemma Delogu, Rosaria Rinaldi

    Alginate-(ALG) and chitosan-(CS) have been extensively used for biomedical applications; however, data relative to immune responses exerted by them are scarce. We synthesized a submicron vesicle system (SV) displaying a CS shell over an ALG core. Intravenous injection of these promising carriers could be a possible route of delivery; therefore, we evaluated their impact on human peripheral blood mononuclear cells (PBMCs). By this ex vivo approach, we established how SV chemical-physical characteristics affected the immune cells in terms of cellular uptake, viability and state of activation. By flow cytometry, we demonstrated that SVs were internalized by PBMCs with differential trends. No substantial necrotic and apoptotic signals were recorded and SVs weakly affected activation status of PBMCs (concerning the markers CD69, CD25, CD80 and the cytokines TNF-α and IL-6), showing high immune biocompatibility and low immunomodulating properties. Our findings gain particular value towards the biomedical applications of SVs, and make these polymer-based structures more attractive for translation into clinical uses.

    更新日期:2018-07-16
  • 更新日期:2018-07-16
  • Short oligopeptides with three cystein residues as models of sulphur-rich Cu(I)- and Hg(II)-binding sites in proteins≈
    Metallomics (IF 4.069) Pub Date : 2018-07-16
    Edit Mesterházy, colette Lebrun, Serge Crouzy, Attila Jancsó, Pascale Delangle

    The essential Cu(I) and the toxic Hg(II) ions possess similar coordination properties and therefore, similar cysteine rich proteins participate in the control of their intracellular concentration. In this work we present the metal binding properties of linear and cyclic model peptides incorporating the three cysteine motifs, CxCxxC or CxCxC, found in metallothioneins. Cu(I) binding to the series of peptides at physiological pH revealed to be rather complicated, with the formation of mixtures of polymetallic species. In contrast, the Hg(II) complexes display well-defined structures with spectroscopic features characteristic for a HgS2 and HgS3 coordination mode at pH = 2.0 and 7.4, respectively. Stability data reflect a ca. 20 orders of magnitude larger affinity of the peptides for Hg(II) (logβHgPpH 7.4 ≈ 41) than for Cu(I) (logβCuPpH 7.4 ≈ 18). The different behaviour with the two metal ions demonstrates that the use of Hg(II) as a probe for Cu(I), coordinated by thiolate ligands in water, may not always be fully appropriate.

    更新日期:2018-07-16
  • Nickel(II)-promoted specific hydrolysis of zinc finger proteins
    Metallomics (IF 4.069) Pub Date : 2018-07-04
    Agnieszka Belczyk-Ciesielska, Brigitta Csipak, Bálint Hajdu, Aleksandra Sparavier, Masamitsu N. Asaka, Kyosuke Nagata, Béla Gyurcsik, Wojciech Bal
    更新日期:2018-07-16
  • A Potent Conformation-Constrained Synthetic Peptide Mimic of a Homeodomain Selectively Regulates Target Genes in Cells
    ACS Chem. Biol. (IF 4.592) Pub Date : 2018-07-16
    Basusree Ghosh, Liberalis Debraj Boila, Susobhan Choudhury, Priya Mondal, Sayan Bhattacharjee, Samir Kumar Pal, Amitava Sengupta, Siddhartha Roy
    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
  • Anti-cancer activity of di- and tri-organotin(IV) compounds with D-(+)-Galacturonic acid on human tumor cells
    J. Inorg. Biochem. (IF 3.063) Pub Date : 2018-04-09
    Alessandro Attanzio, Maristella Ippolito, Maria Assunta Girasolo, Filippo Saiano, Archimede Rotondo, Simona Rubino, Luigi Mondello, Massimo L. Capobianco, Piera Sabatino, Luisa Tesoriere, Girolamo Casella

    We have compared the anti-proliferative activity in vitro, of R2SnGala (1-3) [R = Me, n-Bu, Ph] and novel R3SnGala (4, 5) [R = Me, n-Bu] with D-(+)-Galacturonic acid [HGala; Galaq-, q = (2) and (1) for R2SnGala and R3SnGala, respectively] compounds, towards human tumor cell lines of intestinal carcinoma (HCT-116) and breast adenocarcinoma (MCF-7). The new synthesized 4 and 5 compounds were characterized, in solution, by 1H, 13C and 119Sn NMR, that showed that HGala acts as monoanionic moiety and evidenced the dynamic behavior of the compounds, due to inter-conversions involving the anomeric carbon atom of the ligand. Cell viability, apoptosis induction and cell cycle distribution were analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay and flow cytometry, respectively. The cytotoxicity of the compounds, in the micro-submicromolar range, changed in the order of the organotin(IV) moieties, according to 5 > 3 > 2, while 1 and 4, containing MenSn(IV) (n = 2,3) moieties, were ineffective. Compound 5 showed peculiar cytotoxic effects. It did not cause time dependent inhibition of cell growth nor accumulated into the cells. Cell death induced by the active 2, 3, and 5, was shown to be apoptotic by measuring the exposure of phosphatidylserine to the outer membrane and the loss of mitochondrial potential. All the cytotoxic compounds induced an accumulation of cells in the subG0/G1phase, while only 2 and 3 perturbed the cell cycle confining viable cells in G0/G1phase. Finally, none of the compounds investigated affected the viability of normal intestinal or liver cells, indicating selectivity towards tumor cells.

    更新日期:2018-07-14
  • Structural Lipids Enable the Formation of Functional Oligomers of the Eukaryotic Purine Symporter UapA
    Cell Chem. Bio. (IF 5.592) Pub Date : 2018-04-19
    Euan Pyle, Antreas C. Kalli, Sotiris Amillis, Zoe Hall, Andy M. Lau, Aylin C. Hanyaloglu, George Diallinas, Bernadette Byrne, Argyris Politis
    更新日期:2018-07-14
  • β-Glucan-induced cooperative oligomerization of Dectin-1 C-type lectin-like domain
    Glycobiology (IF 3.664) Pub Date : 2018-04-20
    Hari P Dulal, Yoshiyuki Adachi, Naohito Ohno, Yoshiki Yamaguchi

    Dectin-1 is a C-type lectin-like pattern recognition receptor that recognizes β(1–3)-glucans present on non-self pathogens. It is of great importance in innate immunity to understand the mechanism whereby Dectin-1 senses β(1–3)-glucans and induces intracellular signaling. In this study, we characterize the ligand binding and ligand-induced oligomerization of murine Dectin-1 using its C-type lectin-like domain (CTLD). Interaction of CTLD with laminarin, a β-glucan ligand, induced a tetramer of CTLD, as evidenced by size exclusion chromatography and multi-angle light scattering. Component analysis suggested a stoichiometry of four CTLD molecules bound to four laminarin molecules. Dimers and trimers of CTLD were not detected suggesting cooperative oligomerization. In order to map the amino acid residues of CTLD involved in β-glucan binding and domain oligomerization, we performed site-directed mutagenesis on surface-exposed and most conserved amino acid residues. Among the mutants examined, W221A, H223A and Y228A abolished oligomer formation. Since these residues are spatially arranged to form a hydrophobic groove, it is likely that W221, H223 and Y228 are directly involved in β-glucan binding. Interestingly, mutation of the residues on the other side of the hydrophobic groove, including Y141, R145 and E243, also exhibited reduced oligomer formation, suggesting involvement in protein–protein interactions guided by laminarin. Ligand titration using intrinsic tryptophan fluorescence revealed that wild-type CTLD binds laminarin cooperatively with a Hill coefficient of ~3, while the oligomer-reducing mutations, inside and outside the putative binding site abolish or decrease cooperativity. We suggest that the ligand-induced cooperative oligomer formation of Dectin-1 is physiologically relevant in sensing exogenous β-glucan and triggering intracellular signaling.

    更新日期:2018-07-14
  • Unraveling synthesis of the cryptococcal cell wall and capsule
    Glycobiology (IF 3.664) Pub Date : 2018-04-10
    Zhuo A Wang, Lucy X Li, Tamara L Doering

    Fungal pathogens cause devastating infections in millions of individuals each year, representing a huge but underappreciated burden on human health. One of these, the opportunistic fungus Cryptococcus neoformans, kills hundreds of thousands of patients annually, disproportionately affecting people in resource-limited areas. This yeast is distinguished from other pathogenic fungi by a polysaccharide capsule that is displayed on the cell surface. The capsule consists of two complex polysaccharide polymers: a mannan substituted with xylose and glucuronic acid, and a galactan with galactomannan side chains that bear variable amounts of glucuronic acid and xylose. The cell wall, with which the capsule is associated, is a matrix of alpha and beta glucans, chitin, chitosan, and mannoproteins. In this review, we focus on synthesis of the wall and capsule, both of which are critical for the ability of this microbe to cause disease and are distinct from structures found in either model yeasts or the mammals afflicted by this infection. Significant research effort over the last few decades has been applied to defining the synthetic machinery of these two structures, including nucleotide sugar metabolism and transport, glycosyltransferase activities, polysaccharide export, and assembly and association of structural elements. Discoveries in this area have elucidated fundamental biology and may lead to novel targets for antifungal therapy. In this review, we summarize the progress made in this challenging and fascinating area, and outline future research questions.

    更新日期:2018-07-14
  • Demystifying O-GlcNAcylation: hints from peptide substrates
    Glycobiology (IF 3.664) Pub Date : 2018-03-22
    Jie Shi, Rob Ruijtenbeek, Roland J Pieters

    O-GlcNAcylation, analogous to phosphorylation, is an essential post-translational modification of proteins at Ser/Thr residues with a single β-N-acetylglucosamine moiety. This dynamic protein modification regulates many fundamental cellular processes and its deregulation has been linked to chronic diseases such as cancer, diabetes and neurodegenerative disorders. Reversible attachment and removal of O-GlcNAc is governed only by O-GlcNAc transferase and O-GlcNAcase, respectively. Peptide substrates, derived from natural O-GlcNAcylation targets, function in the catalytic cores of these two enzymes by maintaining interactions between enzyme and substrate, which makes them ideal models for the study of O-GlcNAcylation and deglycosylation. These peptides provide valuable tools for a deeper understanding of O-GlcNAc processing enzymes. By taking advantage of peptide chemistry, recent progress in the study of activity and regulatory mechanisms of these two enzymes has advanced our understanding of their fundamental specificities as well as their potential as therapeutic targets. Hence, this review summarizes the recent achievements on this modification studied at the peptide level, focusing on enzyme activity, enzyme specificity, direct function, site-specific antibodies and peptide substrate-inspired inhibitors.

    更新日期:2018-07-14
  • Multicolored Protein Nanoparticles: Synthesis, Characterization, and Cell Uptake
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-13
    Bobbi S. Stromer, Sonali Roy, Melissa R. Limbacher, Bardwi Narzary, Manobjyoti Bordoloi, Julia Waldman, Challa Vijaya Kumar
    更新日期:2018-07-14
  • Hydrolytically Degradable PEGylated Polyelectrolyte Nanocomplexes for Protein Delivery
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-13
    Alexander K. Andrianov, Alexander Marin, Andre P. Martinez, Jacob L. Weidman, Thomas R. Fuerst
    更新日期:2018-07-14
  • Orientation Control of Trametes Laccases on a Carbon Electrode Surface to Understand the Orientation Effect on the Electrocatalytic Activity
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-12
    Yasuto Hoshikawa, Alberto Castro-Muñiz, Hanako Tawata, Kouichi Nozaki, Shohei Yamane, Tetsuji Itoh, Takashi Kyotani
    更新日期:2018-07-14
  • 更新日期:2018-07-14
  • Phase Behavior of Acetylated Cellulose Nanocrystals and Origins of the Cross-Hatch Birefringent Texture
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-12
    Mingzhe Jiang, Matthew F. McMillan, Virginia Davis, Christopher L Kitchens
    更新日期:2018-07-14
  • Secondary-Structure-Mediated Hierarchy and Mechanics in Polyurea–Peptide Hybrids
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-12
    Lindsay E. Matolyak, Chase B. Thompson, Bingrui Li, Jong K. Keum, Jonathan E. Cowen, Richard S. Tomazin, LaShanda T. J. Korley
    更新日期:2018-07-14
  • Characterizing the Surface Coverage of Protein-Gold Nanoparticle Bioconjugates
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-13
    Rachel B. Kozlowski, Ashwin Ragupathi, R. Brian Dyer

    Functional enzyme-nanoparticle bioconjugates are increasingly important in biomedical and biotechnology applications such as drug delivery and biosensing. Optimization of the function of such bioconjugates requires careful control and characterization of their structures and activity, but current methods are inadequate for this purpose. A key shortcoming of existing approaches is the lack of an accurate method for quantitating protein content of bioconjugates for low (monolayer) surface coverages. In this study, an integrated characterization methodology for protein-gold nanoparticle (AuNP) bioconjugates is developed, with a focus on site-specific attachment and surface coverage of protein on AuNPs. Single cysteine containing mutants of dihydrofolate reductase are covalently attached to AuNPs with diameters of 5, 15 and 30 nm, providing a range of surface curvature. Site-specific attachment to different regions of the protein surface is investigated, including attachment to a flexible loop versus a rigid alpha helix. Characterization methods include SDS-PAGE, UV/Vis spectrophotometry, Dynamic Light Scattering, and a novel fluorescence-based method for accurate determination of low protein concentration on AuNPs. An accurate determination of both protein and AuNP concentration in conjugate samples allows for the calculation of the surface coverage. We find that surface coverage is related to the surface curvature of the AuNP, with a higher surface coverage observed for higher surface curvature. The combination of these characterization methods is important for understanding the functionality of protein-AuNP bioconjugates, particularly enzyme activity.

    更新日期:2018-07-14
  • One-Pot Synthesis of pH/Redox Responsive Polymeric Prodrug and Fabrication of Shell Cross-Linked Prodrug Micelles for Antitumor Drug Transportation
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-13
    Lei Li, Dian Li, Mingzu Zhang, Jinlin He, Jian Liu, Peihong Ni

    Shell cross-linked (SCL) polymeric prodrug micelles have the advantages of good blood circulation stability and high drug content. Herein, we report on a new kind of pH/redox responsive dynamic covalent SCL micelles, which was fabricated by self-assembly of a multifunctional polymeric prodrug. At first, a macroinitiator PBYP-ss-iBuBr was prepared via ring-opening polymerization (ROP), wherein PBYP represents poly[2-(but-3-yn-1-yloxy)-2-oxo-1,3,2-dioxaphospholane]. Subsequently, PBYP-hyd-DOX-ss-P(DMAEMA-co-FBEMA) prodrug was synthesized by one-pot method with a combination of atom transfer radical polymerization (ATRP) and Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reaction using a doxorubicin (DOX) derivative containing azide group to react with the alkynyl group of the side chain in the PBYP block, while DMAEMA and FBEMA are the abbriviations of N,N-(2-dimethylamino)ethyl methacrylate and 2-(4-formylbenzoyloxy)ethyl methacrylate, respectively. The chemical structures of the polymer precurcors and the prodrugs have been fully characterized. The SCL prodrug micelles were obtained by self-assembly of the prodrug and adding cross-linker dithiol bis(propanoic dihydrazide) (DTP). Compared with the shell uncross-linked prodrug micelles, the SCL prodrug micelles can enhance the stability and prevent the drug from leaking in the body during blood circulation. The average size and morphology of the SCL prodrug micelles were measured by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. The SCL micelles can be dissociated under moderately acidic or/and reductive microenvironment, that is, endosomal/lysosomal pH medium or high GSH level in the tumorous cytosol. The results of DOX release aslo confirmed that the SCL prodrug micelles possessed pH/reduction responsive properties. Cytotoxicity and cellular uptake analyses further revealed that the SCL prodrug micelles could be rapidly internalized into tumor cells through endocytosis and efficiently release DOX into the HeLa and HepG2 cells, which could efficient inhibit the cell proliferation. This study provides a fast and precise synthesis method for preparing multifunctional polymer prodrugs, which hold great potential for optimal antitumor therapy.

    更新日期:2018-07-14
  • Fluorescent Sulfonate-Based Inorganic-Organic Hybrid Nanoparticles for Staining and Imaging
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-13
    Marieke Poß, Eva Zittel, Anna Meschkov, Ute Schepers, Claus Feldmann

    Sulfonate-based inorganic-organic hybrid nanoparticles (IOH-NPs) with the general saline composition [Gd(OH)]2+n/2[Rdye(SO3)n]n– showing optical absorption and emission in the blue to red spectral regime are presented for the first time. All IOH-NPs are prepared via straightforward aqueous synthesis and instantaneously result in colloidally highly stable suspensions with mean particle diameters of 40-50 nm and high zeta potentials (–20 to –40 mV at pH 7.0). Specifically, the IOH-NPs comprise [Gd(OH)]2+2[CSB]4–, [Gd(OH)]2+2[DB71]4–, [Gd(OH)]2+[NFR]2–, [Gd(OH)]2+[AR97]2–, and [Gd(OH)]2+2[EB]4– showing blue, orange, red and infrared absorption and emission ([CSB]: Chicago Sky Blue; [DB71]: Direct Blue 71; [NFR]: Nuclear Fast Red; [AR97]: Acid Red 97; [EB]: Evans Blue). The novel IOH-NPs are characterized by electron microscopy, dynamic light scattering, infrared spectroscopy, energy-dispersive X-ray analysis, thermogravimetry, elemental analysis, and fluorescence spectroscopy. In vitro studies based on HeLa and HUVEC cells were exemplarily performed with [Gd(OH)]2+2[EB]4– IOH-NPs and show intense fluorescence and only moderate toxicity at concentrations of 1 to 10 µg/mL. Based on aqueous synthesis, good colloidal stability, absence of severely toxic metals (e.g. Cd2+, Pb2+), use of molecular dyes that are already known for staining in cell biology and histology, extremely high dye load per nanoparticle (70-80 wt-%), and blue to red absorption and fluorescence, the sulfonate-based IOH-NPs can be highly interesting for staining, fluorescence microscopy and optical imaging.

    更新日期:2018-07-14
  • Synthesis of Modular Brush Polymer-Protein Hybrids using Diazotransfer and Copper Click Chemistry
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-12
    Luis A. Navarro, Daniel L. French, Stefan Zauscher

    Proteoglycans are important brush-like biomacromolecules which serve a variety of functions in the human body. While protein-bottlebrush hybrids are promising proteoglycan mimics, many challenges still exist to robustly produce such polymers. In this paper, we report the modular synthesis of protein-brush hybrids containing elastin-like polypeptides (ELP) as model proteins by copper-catalyzed azide-alkyne cycloaddition. We exploit the recently discovered imidazole-1-sulfonyl azide (ISA) in a diazotransfer reaction to introduce an N-terminal azide onto an ELP. Next, we use a click reaction to couple the azido-ELP to an alkyne-terminated amine-rich polymer followed by a second diazotransfer step to produce an azide-rich backbone that serves as a scaffold. Finally, we used a second click reaction to graft alkyne-terminated poly(oligoethylene glycol methacrylate) (POEGMA) bristles to the azide-rich backbone to produce the final protein-bottlebrush hybrid. We demonstrate the effectiveness of this synthetic path at each step through careful characterization with 1H-NMR, FTIR, GPC, and diagnostic test reactions on SDS-PAGE. Final reaction products could be consistently obtained for a variety of different molecular weight backbones with final total grafting efficiencies around 70 %. The high-yielding reactions employed in this highly modular approach allow for the synthesis of protein-bottlebrush hybrids with different proteins and brush polymers. Additionally, the mild reaction conditions used have the potential to avoid damage to proteins during synthesis.

    更新日期:2018-07-14
  • Bioinspired pH- and Temperature-Responsive Injectable Adhesive Hydrogels with Polyplexes Promotes Skin Wound Healing
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-13
    Thai Minh Duy Le, Huu Thuy Trang Duong, Thavasyappan Thambi, V.H. Giang Phan, Ji Hoon Jeong, Doo Sung Lee

    Despite great potential, the delivery of genetic materials into cells or tissues of interest remains challenging owing to their susceptibility to nuclease degradation, lack of permeability to the cell membrane, and short in vivo half-life, which severely restrict their widespread use in therapeutics. To surmount these shortcomings, we developed a bioinspired in situ-forming pH- and temperature-sensitive injectable hydrogel depot that could control the delivery of DNA-bearing polyplexes for versatile biomedical applications. A series of multiblock copolymer, comprised of water soluble poly(ethylene glycol) (PEG) and pH- and temperature-responsive poly(sulfamethazine ester urethane) (PSMEU), has been synthesized as in situ-forming injectable hydrogelators. The free-flowing PEG-PSMEU copolymer sols at high pH and room temperature (pH 8.5, 23 oC) were transformed to stable gel at the body condition (pH 7.4, 37 oC). Physical and mechanical properties of hydrogels, including their degradation rate and viscosity, are elegantly controlled by varying the composition of urethane ester units. Subcutaneous administration of free-flowing PEG-PSMEU copolymer sols to the dorsal region of Sprague-Dawley rats instantly formed hydrogel depot. The degradation of the hydrogel depot was slow at the beginning and found to be bioresorbable after two months. Cationic protein or DNA-bearing polyplex-loaded PEG-PSMEU copolymer sols formed stable gel and controlled its release over 10 days in vivo. Owing to the presence of urethane linkages, the PEG-PSMEU possesses excellent adhesion strength to wide range of surfaces including glass, plastic and fresh organs. More importantly, the hydrogels effectively adhered on human skin and peeled easily without eliciting an inflammatory response. Subcutaneous implantation of PEG-PSMEU copolymer sols effectively sealed the ruptured skin, which accelerated the wound healing process as observed by the skin appendage morphogenesis. The bioinspired in situ-forming pH- and temperature-sensitive injectable adhesive hydrogel may provide a promising platform for myriad biomedical applications as controlled delivery vehicle, adhesive and tissue regeneration.

    更新日期:2018-07-14
  • Tripeptide-stabilized oil-in-water nanoemulsion of an oleic acids-platinum (II) conjugate as an anticancer nanomedicine.
    Bioconjugate Chem. (IF 4.485) Pub Date : 2018-07-12
    Sylwia A Dragulska, Ying Chen, Marek T Wlodarczyk, Mina Poursharifi, Peter Dottino, Rein V. Ulijn, John A Martignetti, Aneta Joanna Mieszawska

    We report a nanoemulsion (NE) which is stabilized by self-assembling tripeptide lysine-tyrosine-phenylalanine (KYF) and encapsulates an oleic acids-platinum conjugate formed using a simple Pt (II) coordination chemistry. The KYF-Pt-NE is evaluated both in cultured ovarian cancer cells and in an in vivo preclinical cancer model and shows pH dependent Pt (II) release, which is low at physiological pH and enhanced at tumoral pH. The biological activity of KYF-Pt-NE, evaluated in multiple ovarian cancer cell lines, is significantly higher when compared to analogous Pt (II) complex used in the clinic. Concurrently, the KYF-Pt-NE platform shows good compatibility with the immune system. Preliminary in vivo testing of KYF-Pt-NE with tumor bearing mice indicate efficient Pt (II) delivery to the tumor. Together, these results demonstrate the potential of peptide-stabilized nanoemulsions, specifically KYF-Pt-NE as an effective nanomedicine against cancer.

    更新日期:2018-07-14
  • A top-down approach to produce protein-functionalized and highly thermally stable cellulose fibrils
    Biomacromolecules (IF 5.738) Pub Date : 2018-07-13
    Franck Quero, Genesis Opazo, Yadong Zhao, Aymeric Feschotte-Parazon, Jeimy Fernández, Abraham Mackenna, Marcos Flores

    Protein-functionalized cellulose fibrils, having various amounts of covalently bonded proteins at their surface, were successfully extracted from the tunic of Pyura Chilensis tunicates using successive alkaline extractions. Pure cellulose fibrils were also obtained by further bleaching and were used as reference material. Extraction yields of protein-functionalized cellulose fibrils were within the range of 62-76% by weight based on the dry initial tunic powder. Fourier-transform infrared and Raman spectroscopy confirmed the preservation of residual protein at the surface of cellulose fibrils, which was then quantified by X-ray photoelectron spectroscopy. The protein-functionalized cellulose fibrils were found to have relatively high crystallinity and their cellulose I crystalline structure was preserved upon applying alkaline treatments. The extracted cellulosic materials were found to be constituted of fibrils having a ribbon-like morphology with widths ranging from ~30 nm up to ~400 nm. These protein-functionalized cellulose fibrils were found to have outstanding thermal stability with one of them having onset and peak degradation temperatures of ~350 °C and 374 °C, respectively. These values were found to be 24 and 41 °C higher than for bleached cellulose.

    更新日期:2018-07-14
  • Mechanism of Substrate Recognition and Catalysis of the Haloalkanoic Acid Dehalogenase Family Member α-Phosphoglucomutase
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Chunchun Zhang, Karen N. Allen, Debra Dunaway-Mariano
    更新日期:2018-07-14
  • Effects of Disease-Causing Mutations on the Conformation of Human Apolipoprotein A-I in Model Lipoproteins
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Christopher J. Wilson, Madhurima Das, Shobini Jayaraman, Olga Gursky, John R. Engen
    更新日期:2018-07-14
  • Peroxidase versus Peroxygenase Activity: Substrate Substituent Effects as Modulators of Enzyme Function in the Multifunctional Catalytic Globin Dehaloperoxidase
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Ashlyn H. McGuire, Leiah M. Carey, Vesna de Serrano, Safaa Dali, Reza A. Ghiladi
    更新日期:2018-07-14
  • The Methanosarcina mazei MM2060 Gene Encodes a Bifunctional Kinase/Decarboxylase Enzyme Involved in Cobamide Biosynthesis
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Norbert K. Tavares, Carmen L. Zayas, Jorge C. Escalante-Semerena
    更新日期:2018-07-14
  • d-Amino Acid-Mediated Translation Arrest Is Modulated by the Identity of the Incoming Aminoacyl-tRNA
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Rachel C. Fleisher, Virginia W. Cornish, Ruben L. Gonzalez
    更新日期:2018-07-14
  • C-Methylation Catalyzed by Fom3, a Cobalamin-Dependent Radical S-adenosyl-l-methionine Enzyme in Fosfomycin Biosynthesis, Proceeds with Inversion of Configuration
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Shusuke Sato, Fumitaka Kudo, Tomohisa Kuzuyama, Friedrich Hammerschmidt, Tadashi Eguchi
    更新日期:2018-07-14
  • Impact of D1-V185 on the Water Molecules That Facilitate O2 Formation by the Catalytic Mn4CaO5 Cluster in Photosystem II
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Christopher J. Kim, Han Bao, Robert L. Burnap, Richard J. Debus
    更新日期:2018-07-14
  • 更新日期:2018-07-14
  • 更新日期:2018-07-14
  • Characterization and Use of TurboLuc Luciferase as a Reporter for High-Throughput Assays
    Biochemistry (IF 2.997) Pub Date : 2018-04-20
    Douglas S. Auld, Janaki Narahari, Pei-i Ho, Dominick Casalena, Vy Nguyen, Evelina Cirbaite, Doug Hughes, John Daly, Brian Webb
    更新日期:2018-07-14
  • Evolution of a Thermophilic Strand-Displacing Polymerase Using High-Temperature Isothermal Compartmentalized Self-Replication
    Biochemistry (IF 2.997) Pub Date : 2018-04-18
    John N. Milligan, Raghav Shroff, Daniel J. Garry, Andrew D. Ellington
    更新日期:2018-07-14
  • Watching Three-Dimensional Movements of Single Membrane Proteins in Lipid Bilayers
    Biochemistry (IF 2.997) Pub Date : 2018-04-12
    Li Ma, Ying Li, Jianbing Ma, Shuxin Hu, Ming Li
    更新日期:2018-07-14
  • Single-Molecule Fluorescence Detection of the Epidermal Growth Factor Receptor in Membrane Discs
    Biochemistry (IF 2.997) Pub Date : 2018-04-06
    Steven D. Quinn, Shwetha Srinivasan, Jesse B. Gordon, Wei He, Kermit L. Carraway, Matthew A. Coleman, Gabriela S. Schlau-Cohen
    更新日期:2018-07-14
  • Changes in Protein Dynamics in Escherichia coli SufS Reveal a Possible Conserved Regulatory Mechanism in Type II Cysteine Desulfurase Systems
    Biochemistry (IF 2.997) Pub Date : 2018-04-05
    Dokyong Kim, Harsimran Singh, Yuyuan Dai, Guangchao Dong, Laura S. Busenlehner, F. Wayne Outten, Patrick A. Frantom
    更新日期:2018-07-14
  • Bacterial Model Membranes Reshape Fibrillation of a Functional Amyloid Protein
    Biochemistry (IF 2.997) Pub Date : 2018-04-02
    Ravit Malishev, Razan Abbasi, Raz Jelinek, Liraz Chai
    更新日期:2018-07-14
  • An open library of human kinase domain constructs for automated bacterial expression
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Steven K. Albanese, Daniel L. Parton, Mehtap Isik, Lucelenie Rodríguez-Laureano, Sonya M Hanson, Julie M. Behr, Scott Gradia, Chris Jeans, Nicholas M. Levinson, Markus A Seeliger, John Damon Chodera

    Kinases play a critical role in many cellular signaling pathways and are dysregulated in a number of diseases, such as cancer, diabetes, and neurodegeneration. Since the FDA approval of imatinib in 2001, therapeutics targeting kinases now account for roughly 50% of current cancer drug discovery efforts. The ability to explore human kinase biochemistry, biophysics, and structural biology in the laboratory is essential to making rapid progress in understanding kinase regulation, designing selective inhibitors, and studying the emergence of drug resistance. While insect and mammalian expression systems are frequently used for the expression of human kinases, bacterial expression systems are superior in terms of simplicity and cost-effectiveness but have historically struggled with human kinase expression. Following the discovery that phosphatase coexpression could produce high yields of Src and Abl kinase domains in bacterial expression systems, we have generated a library of 52 His-tagged human kinase domain constructs that express above 2 µg/mL culture in a simple automated bacterial expression system utilizing phosphatase coexpression (YopH for Tyr kinases, Lambda for Ser/Thr kinases). Here, we report a structural bioinformatics approach to identify kinase domain constructs previously expressed in bacteria likely to express well in a simple high-throughput protocol, experiments demonstrating our simple construct selection strategy selects constructs with good expression yields in a test of 84 potential kinase domain boundaries for Abl, and yields from a high-throughput expression screen of 96 human kinase constructs. Using a fluorescence-based thermostability assay and a fluorescent ATP-competitive inhibitor, we show that the highest-expressing kinases are folded and have well-formed ATP binding sites. We also demonstrate how the resulting expressing constructs can be used for the biophysical and biochemical study of clinical mutations by engineering a panel of 48 Src mutations and 46 Abl mutations via single-primer mutagenesis and screening the resulting library for expression yields. The wild-type kinase construct library is available publicly via Addgene, and should prove to be of high utility for experiments focused on drug discovery and the emergence of drug resistance.

    更新日期:2018-07-14
  • Open and Closed Form of Maltose Binding Protein in Its Native and Molten Globule State as Studied by EPR Spectroscopy
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Benjamin Selmke, Peter P Borbat, Chen Nickolaus, Raghavan Varadarajan, Jack H. Freed, Wolfgang E. Trommer

    An intensively investigated intermediate state of protein folding is the molten globule state (MG), which contains secondary but hardly any tertiary structure. In previous work we have determined the distances between interacting spins within maltose binding protein (MBP) in its native state using continuous wave and double electron-electron resonance (DEER) EPR spectroscopy. Seven double mutants, MBP01 to MBP07, had been employed to investigate the structure within the two domains of MBP. DEER data nicely corroborated the previously available X-ray data. Even in its MG state, MBP is known to still bind its ligand maltose. We therefore hypothesized that there must be a defined structure around the binding pocket of MBP already in the absence of tertiary structure. To investigate the difference between native and MG state in open and closed form mutants MBP 09-11 were employed. In these the MTS positions were placed near the active site. The complete set of MBP mutants was analyzed at pH 3.2 (MG) and pH 7.4 (native state) using double quantum coherence (DQC) EPR. The values were compared with theoretical predictions of distances between the labels in biradicals constructed by molecular modeling from the crystal structures of MBP in open and closed form and were found to be in excellent agreement. Measurements show a defined structure around the binding pocket of MBP in MG, which explains maltose binding. In both states ligand-free MBP can be found in open and closed form, while ligand-bound MBP only appears in closed form due to maltose binding.

    更新日期:2018-07-14
  • Molecular determinants of substrate specificity in human insulin-degrading enzyme
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    Lazaros Stefanidis, Nicholas D. Fusco, Samantha E. Cooper, Jillian Smith-Carpenter, Benjamin J. Alper

    Insulin-degrading enzyme (IDE) is a 110 kDa chambered zinc metalloendopeptidase that degrades insulin, amyloid beta, and other intermediate-sized aggregation prone peptides that adopt β-structures. Structural studies of IDE in complex with multiple physiological substrates have suggested a role for hydrophobic and aromatic residues of the IDE active site in substrate binding and catalysis. Here, we examine functional requirements for conserved hydrophobic and aromatic IDE active site residues that are positioned within 4.5 Angstroms of IDE bound insulin B chain and amyloid beta peptides in the reported crystal structures for the respective enzyme-substrate complexes. Charge, size, hydrophobicity, aromaticity, and other functional group requirements for substrate binding IDE active site residues were examined through mutational analysis of the recombinant human enzyme and enzyme kinetic studies conducted using native and fluorogenic derivatives of human insulin and amyloid beta peptides. A functional requirement for IDE active site residues F115, A140, F141, Y150, W199, F202, F820, and Y831 was established, and specific contributions of residue charge, size and hydrophobicity in substrate binding, specificity, and proteolysis were demonstrated. IDE mutant alleles that exhibited enhanced or diminished proteolytic activity towards insulin or amyloid beta peptides and derivative substrates were identified.

    更新日期:2018-07-14
  • Photoaffinity cross-linking and unnatural amino acid mutagenesis reveal insights into calcitonin gene-related peptide binding to the calcitonin receptor-like receptor/receptor activity-modifying protein 1 (CLR/RAMP1) complex
    Biochemistry (IF 2.997) Pub Date : 2018-07-13
    John Simms, Romez Uddin, Thomas P. Sakmar, Joseph J. Gingell, Michael L. Garelja, Debbie L. Hay, Margaret A. Brimble, Paul W. R. Harris, Christopher Arthur Reynolds, David R. Poyner

    Calcitonin gene-related peptide (CGRP) binds to the complex of the calcitonin receptor-like receptor (CLR) with receptor activity-modifying protein 1 (RAMP1). It remains unclear how CGRP interacts with the transmembrane domain (including the extracellular loops) of this family B receptor. In the current study, a photoaffinity cross linker, p-azido L-phenylalanine (azF) was incorporated into CLR, chiefly in the second extracellular loop (ECL2) using genetic code expansion and unnatural amino acid mutagenesis. The method was optimised to ensure efficient photolysis of azF residues near to the transmembrane bundle of the receptor. A CGRP analogue modified with fluorescein at position 15 was used for detection of uv-induced cross-linking. The methodology was verified by confirming the known contacts of CGRP to the extracellular domain of CLR. Within ECL2, the chief contacts were I284 on the loop itself and L291, at the top of the 5th transmembrane helix (TM5). Minor contacts were noted along the lip of ECL2 between S286 and L290 and also with M223 in TM3 and F349 in TM6. Full length molecular models of the bound receptor complex suggest that CGRP sits at the top of the TM bundle, with Thr6 of the peptide making contacts with L291 and H295. I284 is likely to contact Leu12 and Ala13 of CGRP, and Leu16 of CGRP is at the ECL/extracellular domain (ECD) boundary of CLR. Reduced potency, Emax and affinity of [Leu16Ala]-human alpha CGRP is consistent with this model. Contacts between Thr6 of CGRP and H295 may be particularly important for receptor activation.

    更新日期:2018-07-14
  • Adaptive Multifunctional Supramolecular Assemblies of Glycopeptides Rapidly Enable Morphogenesis
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Jie Zhou, Xuewen Du, Xiaoyi Chen, Bing Xu

    Despite the well-established biophysical principle of adhesion-guided in vitro morphogenesis, there are few single syn-thetic molecular species that can rapidly enable morphogenesis (e.g., a cell monolayer to cell spheroids) in cell culture because adhesion process inherently involves many signals. Here we show the use of adaptive multifunctional supramo-lecular assemblies of glycopeptides, consisting of cell adhesion sequence and saccharide, to induce cell spheroids rapid-ly from a monolayer of cells. Having a general architecture of N-terminal capping, glycosylation, and an integrin bind-ing sequence, the glycopeptides self-assemble to form a dynamic continuum of nanostructures (i.e., from nanoparticles to nanofibers) to affect the interactions of integrins, E-selectin, and cadherins with their natural ligands and to act adaptively according to cellular environment. Such adaptive (i.e., context-dependent) interactions reduce cell-substratum adhesion and enhance intercellular interactions, which rapidly and transiently induce cell spheroids. This work illustrates a new approach that uses supramolecular assemblies of simple glycopeptides as molecular modulators of biophysical condition of cells for understanding and affecting cell functions in the development of supramolecular biomaterials.

    更新日期:2018-07-14
  • Dynamic states of the ligand-free class A G protein-coupled receptor extracellular side
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Qiansen Zhang, Mang Zhou, Lifen Zhao, Hualiang Jiang, Huaiyu Yang

    G protein-coupled receptors (GPCRs) are the largest family of drug targets. The second extracellular loop (ECL2) and extracellular end of the third transmembrane helix (TM3) are basic structural elements of the GPCR ligand binding site. Currently, the disulphide bond between the two conserved cysteines in the ECL2 and TM3 is considered to be a basic GPCR structural feature. This disulphide bond has a significant effect on receptor dynamics and ligand binding. Here, molecular dynamics (MD) simulations and experimental results show that the two cysteines are distant from one another in the highest population conformational state of ligand-free class A GPCRs and do not form a disulphide bond, indicating that the dynamics of the GPCR extracellular side are different from our conventional understanding. These surprising dynamics should have important effects on the drug binding process. Based on the two distinct ligand-free states, we suggest two kinetic processes for ligand binding to GPCRs. These results challenge our commonly held beliefs regarding both GPCR structural features and ligand binding

    更新日期:2018-07-14
  • Nitrosyl Myoglobins and Their Nitrite Precursors: Crystal Structural and QM/MM Theoretical Investigations into Preferred Fe–NO Ligand Orientations in Myoglobin Distal Pockets
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Bing Wang, Yelu Shi, Jesus Tejero, Samantha M Powell, Leonard M Thomas, Mark T. Gladwin, Sruti Shiva, Yong Zhang, George B. Richter-Addo

    The globular dioxygen-binding heme protein myoglobin (Mb) is present in several species. Its interactions with the simple nitrogen oxides, namely nitric oxide (NO) and nitrite, have been known for decades, but the physiological relevance has only recently become more fully appreciated. We previously reported the O-nitrito binding mode of nitrite to ferric horse heart wild-type (wt) MbIII and human hemoglobin. We have expanded on this work and report the interactions of nitrite with wt sperm whale (sw) Mb and its H64A, H64Q and V68A/I107Y mutants whose dissociation constants increase in the order H64Q < wt < V68A/I107Y < H64A. We also report their X-ray crystal structures that reveal the O-nitrito binding mode of nitrite to these derivatives. The MbII-mediated reductions of nitrite to NO and structural data for these wt and mutant MbII-NOs are described. We show that their FeNO orientations vary with distal pocket identity, with the FeNO moieties pointing towards the hydrophobic interiors when the His64 residue is present, but pointing towards the hydrophilic exterior in the absence of this His64 residue. This correlates with the nature of H-bonding to the bound NO ligand (nitrosyl O vs. N atom). Quantum mechanics and hybrid quantum mechanics/molecular mechanics calculations help elucidate the origin of the experimentally preferred NO orientations. In a few cases, the calculations reproduce the experimentally observed orientations only when the whole protein is taken into consideration.

    更新日期:2018-07-14
  • Steady-state kinetics of phytoglobin-catalyzed hydroxylamine reduction to ammonium
    Biochemistry (IF 2.997) Pub Date : 2018-07-12
    Jagannathan Alagurajan, Navjot Singh Athwal, Mark Scott Hargrove

    Phytoglobins are plant hexacoordinate hemoglobins with reversible coordination of a histidine side chain to the ligand binding site of the heme iron. They mediate electron transfer reactions such as nitric oxide scavenging and are particularly efficient at reducing nitrite and hydroxylamine. Previous work with phytoglobins has focused only on single-turnovers of these reactions and have not revealed whether structural features, such as histidine hexacoordination, play a prominent role in the complete catalytic cycle. The present work characterizes steady-state phytoglobin catalysis of hydroxylamine reduction to ammonium using two different chemical reductants. Km and kcat values were measured for rice phytoglobin, horse myoglobin, human neuroglobin, and a rice phytoglobin mutant protein in which the hexacoordinating histidine has been replaced by leucine (Phyt:H73L). The results demonstrate that phytoglobin catalysis driven by benzyl viologen (BV) is limited only by the dissociation rate constant for the distal histidine. This is consistent with the rate limit in single turnover experiments and demonstrates that the kinetics of hydroxylamine binding, and not phytoglobin reduction, ultimately governs the reaction. Catalysis by the other proteins that either lack or have tighter hexacoordination is much slower, suggesting that facile reversibility of the bond between the distal histidine and the heme iron is needed to allow both substrate binding and heme iron reduction. On the other hand, catalysis driven by dithionite is limited by SO2- concentrations and is similar for all of these proteins, suggesting that dithionite is not a good reducing agent for evaluation of the catalytic properties of hemoglobins.

    更新日期:2018-07-14
  • 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
  • Fine-Tuned Protein Production in Methanosarcina acetivorans C2A
    ACS Synth. Biol. (IF 5.316) Pub Date : 2018-07-13
    Ann A. Karim, Daniel R. Gestaut, Maeva Fincker, John C. Ruth, Eric C. Holmes, Wayne Sheu, Alfred M. Spormann
    更新日期:2018-07-14
  • Optical Activation of TrkA Signaling
    ACS Synth. Biol. (IF 5.316) Pub Date : 2018-07-12
    Liting Duan, Jen M. Hope, Shunling Guo, Qunxiang Ong, Amaury François, Luke Kaplan, Grégory Scherrer, Bianxiao Cui
    更新日期:2018-07-14
  • Robustness of a reconstituted Escherichia coli protein translation system analyzed by computational modeling
    ACS Synth. Biol. (IF 5.316) Pub Date : 2018-07-13
    Tomoaki Matsuura, Kazufumi Hosoda, Yoshihiro Shimizu

    Robustness against environmental changes is one of the major features of biological systems, but its origin is not well understood. We recently constructed a large-scale computational model of an Escherichia coli-based reconstituted in vitro translation system that enumerates all protein synthesis processes in detail. Our model synthesizes a formyl-Met-Gly-Gly tripeptide (MGG peptide) from 27 initial molecular components through 968 biochemical reactions. Among the 968 kinetic parameters, 483 are non-zero parameters, and the simulator was used to determine how perturbations of 483 individual reactions affect the complex reaction network. We found that even when the kinetic parameter was changed from 100- to 0.01-fold, 94% of the changes hardly affected the two indicators of reaction dynamics in MGG peptide synthesis, which represent the yield of the MGG peptide and the initial lag-time of the peptide synthesis. Moreover, none of the indicators increased proportionally to these changes: e.g., a 100-fold increase in the kinetic parameter only increased the yield by 2.2-fold at most, indicating the insensitivity of the reaction network to perturbation. Robustness and insensitivity are likely to be a common feature of large-scale biological reaction networks.

    更新日期:2018-07-14
  • Lipidomics Suggests a New Role for Ceramide Synthase in Phagocytosis
    ACS Chem. Biol. (IF 4.592) Pub Date : 2018-07-13
    Divya Pathak, Neelay Mehendale, Shubham Singh, Roop Mallik, Siddhesh S. Kamat
    更新日期:2018-07-14
  • Unique Fluorescent Imaging Probe for Bacterial Surface Localization and Resistant Enzyme Imaging
    ACS Chem. Biol. (IF 4.592) Pub Date : 2018-04-04
    Hui Ling Chan, Linna Lyu, Junxin Aw, Wenmin Zhang, Juan Li, Huang-Hao Yang, Hirohito Hayashi, Shunsuke Chiba, Bengang Xing
    更新日期:2018-07-14
  • Chemoproteomics-Enabled Covalent Ligand Screening Reveals ALDH3A1 as a Lung Cancer Therapy Target
    ACS Chem. Biol. (IF 4.592) Pub Date : 2018-07-13
    Jessica L. Counihan, Amanda L. Wiggenhorn, Kimberly E. Anderson, Daniel K. Nomura

    Chemical genetics is a powerful approach for identifying therapeutically active small-molecules, but identifying the mechanisms of action underlying hit compounds remains challenging. Chemoproteomic platforms have arisen to tackle this challenge and enable rapid mechanistic deconvolution of small-molecule screening hits. Here, we have screened a cysteine-reactive covalent ligand library to identify hit compounds that impair cell survival and proliferation in non-small cell lung carcinoma cells, but not in primary human bronchial epithelial cells. Through this screen, we identified a covalent ligand hit, DKM 3-42 which impaired both in situ and in vivo lung cancer pathogenicity. We used activity-based protein profiling to discover that the primary target of DKM 3-42 was the catalytic cysteine in aldehyde dehydrogenase 3A1 (ALDH3A1). We performed further chemoproteomics-enabled covalent ligand screening directly against ALDH3A1, and identified a more potent and selective lead covalent ligand, EN40, which inhibits ALDH3A1 activity and impairs lung cancer pathogenicity. We show here that ALDH3A1 represents a potentially novel therapeutic target for lung cancers that express ALDH3A1 and put forth two selective ALDH3A1 inhibitors. Overall, we show the utility of combining chemical genetics screening of covalent ligand libraries with chemoproteomic approaches to rapidly identify anti-cancer leads and targets.

    更新日期:2018-07-14
  • Hydrogen Bond Surrogate Stabilization of β-Hairpins
    ACS Chem. Biol. (IF 4.592) Pub Date : 2018-07-13
    Nicholas Sawyer, Paramjit S. Arora

    Peptide secondary and tertiary structure motifs frequently serve as inspiration for the development of protein-protein interaction (PPI) inhibitors. While a wide variety of strategies have been used to stabilize or imitate α-helices, similar strategies for β-sheet stabilization are more limited. Synthetic scaffolds that stabilize reverse turns, individual β-strands, and cross-strand interactions have provided important insights into β-sheet stability and folding. However, these templates occupy regions of the β-sheet that might impact the β-sheet’s ability to bind at a PPI interface. Here we present the hydrogen bond surrogate (HBS) approach for stabilization of β-hairpin peptides. The HBS linkage replaces a cross-strand hydrogen bond with a covalent linkage, conferring significant conformational and pro-teolytic resistance. Importantly, this approach introduces the stabilizing linkage in the buried β-sheet interior, retains all side chains for further functionalization, and allows efficient solid-phase macrocyclization. We anticipate that HBS stabilization of PPI β-sheets will en-hance the development of β-sheet PPI inhibitors and expand the repertoire of druggable PPIs.

    更新日期:2018-07-14
  • Selenium-enriched yeast inhibited β-amyloid production and modulated autophagy in a triple transgenic mouse model of Alzheimer’s Disease
    Metallomics (IF 4.069) Pub Date : 2018-07-13
    Guoli Song, Chen Chen, Qiuyan Wu, Zhanghao Zhang, Rui Zheng, Yao Chen, Shizhen Jia, Jiazuan Ni

    As the most common cause of progressive intellectual failure in elderly humans, Alzheimer's disease (AD) is pathologically featured by amyloid plaques, synaptic loss, and neurofibrillary tangles. The amyloid plaques are mainly aggregates of amyloid β-peptide (Aβ), a primary factor contributing to the pathogenesis of AD. Elimination or reduction of the level of Aβ is considered an important strategy in AD treatment. The pharmacotherapeutic efficacy of Selenium (Se), an essential biological trace element for mammalian species, has been confirmed in a number of experimental models of neurodegenerative diseases. Selenium-enriched yeast (Se-yeast) is commonly used as a nutritional supplement for Se. In this study, we investigated the effects and underlying mechanisms of Se-yeast on Aβ pathology in a 4-month-old triple transgenic mouse model of AD (3xTg-AD mice). The administration of Se-yeast attenuated the deposition of Aβ in the brains of AD mice, which was concomitant with decreased levels of LC3II. The Se-yeast treatment decreased the level of amyloid-protein precursor (APP), downregulated the activity of AMP-activated protein kinase (AMPK) and upregulated the activity of AKT/mTOR/p70S6K. Furthermore, the levels of p62 also significantly decreased, and the cathepsin D levels increased, accompanied by increased turnover of Aβ and APP in Se-yeast-treated AD mice. In addition to decreasing the generation of Aβ, Se-yeast also inhibited the initiation of autophagy by modulating the AMPK/AKT/mTOR/p70S6K signaling pathway and enhanced autophagic clearance, thus reducing the burden of Aβ accumulation in the brains of AD mice. Our results further highlight the potential therapeutic effects of Se-yeast on AD.

    更新日期:2018-07-14
  • Vanadocene dichloride inhibits cell proliferation by targeting Aurora B
    Metallomics (IF 4.069) Pub Date : 2018-07-13
    Tzu-Chia Ting, Meng-Ya Chang, Tzu-Yen Hsu, Wen-Pin Wang, Yi-Jen Hsieh, Chih-Jui Chang

    Vanadocene dichloride (VDC) was shown to exhibit antitumor properties against a wide spectrum of tumor cell lines. Many studies have been done to reveal the bioactivities of VDC and the interaction mechanism of VDC with biological molecules in test tube. One of the bioactivities of VDC is to arrest cell cycle in G2/M phase. However, its underline mechanisms of action and cytotoxicity profile are still not fully understood. HeLa cells were used in this study, and the IC50 of VDC was 8.61 µM with a 24-hour treatment. We used immunofluorescence staining method to analyze the morphology of cells in mitosis stage to elucidate what defects caused cells to arrest in mitosis. The chromosomal misalignment was found to be the major phenotype. One of the proteins responsible for chromosome alignment at metaphase is Aurora B kinase. Results of immunoblotting assay showed that Aurora B kinase activity was inhibited by VDC treatment. More than 50% of the Aurora B activity was inhibited when cells were treated with VDC in the concentration of 6.25 µM. That VDC was able to induce defects of chromosomal alignment in metaphase by inhibiting the activity of Aurora B kinase is an important mechanism of VDC to be developed as an antitumor agent.

    更新日期:2018-07-14
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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