Download or read book Optimization of Protein Bioconjugation Reactions Using Combinatorial Peptide Libraries written by Leah Witus and published by . This book was released on 2012 with total page 190 pages. Available in PDF, EPUB and Kindle. Book excerpt: The covalent attachment of chemical groups to proteins is an important tool for many areas of chemical biology, including the study of protein function and the creation of protein-based materials. In order to create well-defined protein bioconjugates, methods of site-specfic protein modification are required. We have developed a high-throughput method of optimizing protein bioconjugation reactions using a combinatorial peptide library in which short peptides serve as a model for protein reactivity. The library screening was achieved using a one-bead-one-compound peptide library, a colorimetric detection scheme to identify high-yielding sequences, and a deconvolution method using a built-in peptide truncation ladder for rapid mass spectrometry sequencing of hit beads. We applied this screening platform in the context of a pyridoxal 5'-phosphate (PLP)-mediated transamination reaction specific for the N-terminus, and were able to identify an N-terminal sequence with high reactivity towards this reaction that can be incorporated into proteins of interest to result in high levels of modification. The screening platform was also used for the discovery and optimization of new N-terminal protein modification reactions.
Download or read book Combinatorial Peptide Library Protocols written by Shmuel Cabilly and published by Springer Science & Business Media. This book was released on 2008-02-02 with total page 320 pages. Available in PDF, EPUB and Kindle. Book excerpt: During the course of evolution, an imbalance was created between the rate of vertebrate genetic adaptation and that of the lower forms of living organisms, such as bacteria and viruses. This imbalance has given the latter the advantage of generating, relatively quickly, molecules with unexpected structures and features that carry a threat to vertebrates. To compensate for their weakness, vertebrates have accelerated their own evolutionary processes, not at the level of whole organism, but in specialized cells containing the genes that code for antibody molecules or for T-cell receptors. That is, when an immediate requirement for molecules capable of specific interactions arose, nature has preferred to speed up the mode of Darwinian evolution in pref- ence to any other approach (such as the use of X-ray diffraction studies and computergraphic analysis). Recently, Darwinian rules have been adapted for test tube research, and the concept of selecting molecules having particular characteristics from r- dom pools has been realized in the form of various chemical and biological combinatorial libraries. While working with these libraries, we noticed the interesting fact that when combinatorial libraries of oligopeptides were allowed to interact with different selector proteins, only the actual binding sites of these proteins showed binding properties, whereas the rest of the p- tein surface seemed "inert. " This seemingly common feature of protein- having no extra potential binding sites--was probably selected during evolution in order to minimize nonspecific interactions with the surrounding milieu.
Download or read book Bioconjugate Techniques written by Greg T. Hermanson and published by Elsevier. This book was released on 1996-01-15 with total page 813 pages. Available in PDF, EPUB and Kindle. Book excerpt: Bioconjugate Techniques is the essential guide to the modification and crosslinking of biomolecules for use in research, diagnostics, and therapeutics. It provides highly detailed information on the chemistry, reagent systems, and practical applications for creating labeled or conjugate molecules. It also describes dozens of reactions with details on hundreds of commercially available reagents and the use of these reagents for modifying or crosslinking peptides and proteins, sugars and polysaccharides, nucleic acids and oligonucleotides, lipids, and synthetic polymers. Armed with this information and the abundant protocols provided, readers will form unique complexes that can be used for detecting, quantifying, and targeting important analytes. This book helps readers make: high activity antibody-enzymes conjugates, immunotoxins, immunogen complexes, liposome conjugates; as well as biotinylated molecules, avidin or streptavidin conjugates, colloidal gold labeled proteins, PEG or dextran complexes, labeled oligonucleotide probes, and fluorescently tagged or radiolabeled molecules. This book is the first to thoroughly capture the entire field of bioconjugate chemistry in a single volume Serves as a practical guide to modification and cross-linking technology for research, diagnostics, and therapeutics Provides useful, detailed, easy-to-follow, step-by-step protocols Contains easy-to-read, and easy-to-understand key concepts for making bioconjugates of all types Efficiently covers the chemistry of bioconjugation, the major reagents available for modification and cross-linking, and the application of these reagents to the synthesis of highly active conjugates Cites over more than references keyed to concepts covered in the book Uses more than 600 figures to illustrate bioconjugate reagents, their reactions, and applications Suggests sources for all key reagents
Download or read book Development of Oxidative Bioconjugation Methodology for the Site Selective Modification of the Electron Rich Aromatic Amino Acids written by Kristen Lee Seim and published by . This book was released on 2013 with total page 198 pages. Available in PDF, EPUB and Kindle. Book excerpt: As applications of protein-based materials become increasingly complex, there is a growing need for an expanded set of bioconjugation reactions that can attach synthetic components to defined locations on protein substrates. Although the introduction of artificial amino acids is a useful method for achieving site-selective protein modification, there are advantages to targeting native amino acids that can be introduced without specialized expression systems. This work describes the discovery, development, and application of new oxidation-based bioconjugation methods that target the native electron-rich aromatic amino acids, tyrosine and tryptophan. These methods include a peptide modification strategy that uses synthetic reagents to site-selectively introduce and modify oxidized tryptophan residues, as well as series of protein modification strategies that use cerium(IV) ammonium nitrate (CAN) to oxidatively couple tyrosine and tryptophan residues to electron rich aniline derivatives. One of these reactions, a CAN-mediated oxidative coupling between anisidine derivatives and tyrosine residues, has been applied towards the synthesis of a novel protein-based material and the site-selectivity of this reaction has been explored using combinatorial peptides libraries. Overall, these unprecedented modification strategies target these under-utilized amino acids with excellent chemoselectivity, greatly augmenting the important and growing list of useful bioconjugation techniques.
Download or read book Synthetic Combinatorial Peptide Libraries and Their Application in Decoding Biological Interactions written by Michael Cameron Sweeney and published by . This book was released on 2005 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Abstract: The synthesis of peptides was revolutionized by the adoption of solid-phase synthetic techniques. Subsequent improvement, evolution, and refinement of this chemical technique has allowed research into areas of biology not previously accessible with such speed and breadth. Because of the efficiency and flexibility of the chemistry involved in peptide synthesis, libraries representing millions of unique natural, modified, or unnatural peptides can be constructed rapidly and in high enough purity as to obviate the need for purification. In this work, libraries were synthesized for screening against individual protein domains in an effort to both determine the preferred peptidyl binding partner types for each, as well as to establish an optimized, broadly applicable methodology for screening other domains. One of the problems encountered during the development of the screening methodology was the low success-rate of sequence determination for the peptides selected by each domain. Herein we report the successful modification of the peptide ladder mass spectrometry sequencing technique referred to as partial Edman degradation (PED). Success-rates were improved to greater than 90% for full-length sequencing determination of peptide up to 8-mers, even for more difficult phosphotyrosine (pY)-containing peptides. As a result of this improvement, three pY-binding Src Homology 2 (SH2) domains and two N-terminus binding Baculoviral Inhibitor-of-Apoptosis Repeat (BIR) domains were screened against their respective libraries and the preferred ligand types for each was determined. The advantage of sequencing by the PED method became especially clear in the case of the N-terminal SH2 (N-SH2) domain of Src Homology 2 Protien Tyrosine Phosphatase 2 (SHP-2) as previously unidentified sub-classes of binding consensus motifs were distinguishable due to the discreet nature of the sequencing technique. This work demonstrates the usefulness and potential generality of peptide library screening by this method.
Download or read book Chemical Approaches to the Synthesis of Peptides and Proteins written by Paul Lloyd-Williams and published by CRC Press. This book was released on 1997-04-23 with total page 306 pages. Available in PDF, EPUB and Kindle. Book excerpt: Organic chemists working on the synthesis of natural products have long found a special challenge in the preparation of peptides and proteins. However, more reliable, more efficient synthetic preparation methods have been developed in recent years. This reference evaluates the most important synthesis methods available today, and also considers methods that show promise for future applications. This text describes the state of the art in efficient synthetic methods for the synthesis of both natural and artificial large peptide and protein molecules. Subjects include an introduction to basic topics, linear solid-phase synthesis of peptides, peptide synthesis in solution, convergent solid-phase synthesis, methods for the synthesis of branched peptides, formation of disulfide bridges, and more. The book emphasizes strategies and tactics that must be considered for the successful synthesis of peptides.
Download or read book Bioconjugation Protocols written by Christof M. Niemeyer and published by Springer Science & Business Media. This book was released on 2008-02-04 with total page 329 pages. Available in PDF, EPUB and Kindle. Book excerpt: There are a number of outstanding volumes that provide a comprehensive overview of bioconjugation techniques. However, many of the conventional approaches to the synthesis of chemically modified protein conjugates lack efficient means to control the stoichiometry of conjugation, as well as the s- cific site of attachment of the conjugated moiety. Moreover, the recent dev- opments in microarray technologies as well as in nanobiotechnology—a novel field of research rapidly evolving at the crossroads of physics, chemistry, b- technology, and materials science—call for a summary of modern bioconjugation strategies to overcome the limitations of the classical approaches. Bioconjugation Protocols: Methods and Strategies is intended to provide an update of many of the classic techniques and also to introduce and summarize newer approaches that go beyond the pure biomedical applications of bioconjugation. The purpose of Bioconjugation Protocols: Methods and Str- egies is therefore to provide instruction and inspiration for all those scientists confronting the challenges of semisynthesizing functional biomolecular reagents for a wide variety of applications ranging from novel biomedical diagnostics, to therapeutics, to biomaterials. Part I contains seven protocols for the preparation of protein conjugates.
Download or read book Development and Optimization of Bioconjugations to Probe and Modulate Protein Function written by Christopher Travis and published by . This book was released on 2018 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Bioconjugate chemistry is a critical field with widespread applications to the visualization, diagnosis, and treatment of various diseases. Thus, it is crucial to investigate and optimize present bioconjugation methods, while continuing to develop novel bioconjugations to expand the scope of the field and provide numerous chemical tools for various applications. This thesis describes the development and optimization of bioconjugations using unnatural amino acid (UAA) technology to prepare homogenous, well-defined macromolecular complexes. First, the utilization of the Glaser-Hay bioconjugation to modulate protein function will be discussed. Next, an investigation into the aqueous Glaser-Hay reaction mechanism and subsequent optimization of the bioconjugation will be presented. A novel [2 + 2 + 2] cycloaddition bioconjugation reaction will then be presented, followed by efforts to generate a multivalent bioconjugate. Finally, efforts to assay and modulate the activity of Cas9 will be examined. This thesis aims to extend the chemical toolbox to probe and control biological systems, with applications in the fields of medicine and pharmaceuticals.
Download or read book New Chromophores and Chromatography Methods for Protein Bioconjugation Reactions written by Trung Nguyẽ̂n and published by . This book was released on 2006 with total page 248 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Development of a Reactive Peptide Sequence for Site selective Bioconjugation written by Suan Lian Tuang and published by . This book was released on 2020 with total page 257 pages. Available in PDF, EPUB and Kindle. Book excerpt: Achieving catalyst-free, site-selective modification of proteins in water is a significant challenge in chemical biology. Issues of residue specificity, site-selectivity, reagent stability, and reaction rate are pervasive in this field, and despite advances over the past few decades, achieving fast, pinpoint modifications of complex molecules remains a tremendous obstacle. Herein, we describe the development of a nine-amino acid motif (Met-Cys-Pro-Phe-Leu-Pro-Val-Val-Tyr) termed engineered reaction (EnAct) tag. The EnAct interface, discovered by iterative screening of peptide libraries, consists of a reactive peptide (EnAct tag) that undergoes rapid (second-order rate constant, k ~ 150 M−1 s−1) nucleophilic aromatic substitution with a perfluoroarene-containing peptide (EnAct probe). Bioconjugation reactions centered on peptide interfaces are emerging as promising strategies to prepare homogeneous biological conjugates, and our results with the EnAct interface represent a 210-fold increase in reaction rate over the previous standard for this class of cysteine arylation. Furthermore, the EnAct sequence consists of all-natural amino acids and thus enables the facile genetic engineering of the sequence onto proteins of interest. We disclose here the incorporation of the EnAct sequence at the C-termini of the IgG antibody trastuzumab heavy chains, which were subsequently conjugated to the EnAct probe with excellent site-selectivity, despite the 32 other Cys residues on this protein. Remarkably, this system's rapid kinetics enabled quantitative conversion in 1.5 hours and at lower substrate concentrations. Finally, this bioconjugation reaction is still selective even in the complex environments of cell lysate mixtures, illustrating the enhanced selectivity and rapid reactivity of the EnAct interface. The appreciable increase in cysteine arylation rate and selectivity achieved with the EnAct sequenced represents a new standard for site-selective bioconjugations using peptide interfaces. To explore the versatility of the reactive peptide sequence, we found that this reactive peptide enabled aqueous arylations of Cys with small molecule electrophiles in mostly water, which was not previously accessed with this class of electrophiles. Furthermore, the perfluoroarene on the probe was found not only to function as an electrophile for thiol arylation, but also to offer a handle for easy elimination to form dehydroalanine. Thus, the EnAct system represents a powerful, versatile, and selective bioconjugation method.
Download or read book Development and Application of Oxidative Coupling Bioconjugation Reactions with Ortho Aminophenols written by Allie Obermeyer and published by . This book was released on 2013 with total page 108 pages. Available in PDF, EPUB and Kindle. Book excerpt: The synthetic modification of proteins plays an important role in the fields of chemical biology and biomaterials science. As applications of protein-based materials continue to become more complex, improved methods for the covalent modification of proteins are needed. Although many methods for the modification of native and artificial amino acids exist, they often require long reaction times or lengthy syntheses of reactive substrates. This work describes the development and application of a suite of bioconjugation reactions that utilize ortho-aminophenols. The oxidative coupling of aniline residues with o-aminophenol substrates was optimized. Potassium ferricyanide was identified as an alternative, mild oxidant for this coupling. These new conditions enabled the use of the oxidative coupling reaction in the presence of free cysteines and glycoslated substrates. Aminophenols were also discovered to react with native residues on protein substrates in addition to artificial aniline moieties. Cysteine and the N-terminus were identified as the reactive residues. The oxidative coupling of o-aminophenols with the N-terminus was optimized to achieve high levels of modification on peptide and protein substrates. The oxidative coupling of anilines and o-aminophenols was applied to the synthesis of a targeted, virus-like particle and to the detection of protein tyrosine-nitration. Overall, these updated and novel oxidative coupling methods expand the utility ortho-aminophenols for the modification of proteins.
Download or read book Bioconjugation written by Mohammed Aslam and published by Stockton Press. This book was released on 1998 with total page 854 pages. Available in PDF, EPUB and Kindle. Book excerpt: Written by two leading experts in the field, Bioconjugation offers invaluable guidance for the design of protein conjugates of all types, covering the coupling of proteins to organic molecules, nucleic acids and solid phases as well as other proteins. Its comprehensive coverage saves time by bringing together information previously available only form a wide variety of sources. An essential reference for college and university libraries supporting research in biochemistry, immunology and medicine.
Download or read book Peptide Conjugation written by Waleed M. Hussein and published by Humana. This book was released on 2022-08-28 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume explores diverse protocols for peptide conjugation, and provides thoroughly tested and scientifically valid techniques that allow researchers and scientists to prepare, purify, characterize, and use peptide conjugation methods for chemical, biochemical, and biological studies. Some of the topics discussed in this book are gold nanoparticles, proteins, pegylated lipids, and vitamins. Chapters also cover enzymatic ligation using sortase A, construction of a phage-displayed cyclic-peptide library, quantum dot-peptide conjugates, and preparation of lipopeptides by CLipPA chemistry. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and comprehensive, Peptide Conjugation: Methods and Protocols is a valuable resource for experienced researches and undergraduate students alike who are interested in learning more about this exciting and developing field.
Download or read book Chemoselective and Bioorthogonal Ligation Reactions written by W. Russ Algar and published by John Wiley & Sons. This book was released on 2017-03-17 with total page 923 pages. Available in PDF, EPUB and Kindle. Book excerpt: This timely, one-stop reference is the first on an emerging and interdisciplinary topic. Covering both established and recently developed ligation chemistries, the book is divided into two didactic parts: a section that focuses on the details of bioorthogonal and chemoselective ligation reactions at the level of fundamental organic chemistry, and a section that focuses on applications, particularly in the areas of chemical biology, biomaterials, and bioanalysis, highlighting the capabilities and benefits of the ligation reactions. With chapters authored by outstanding scientists who range from trailblazers in the field to young and emerging leaders, this book on a highly interdisciplinary topic will be of great interest for biochemists, biologists, materials scientists, pharmaceutical chemists, organic chemists, and many others.
Download or read book Peptide and Protein Bioconjugation written by Philip Anthony Cistrone and published by . This book was released on 2019 with total page 464 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Dynamic Combinatorial Chemistry written by Joost N. H. Reek and published by John Wiley & Sons. This book was released on 2010-02-02 with total page 211 pages. Available in PDF, EPUB and Kindle. Book excerpt: This long-awaited first book on this exciting new field in organic and supramolecular chemistry explains the fundamentals as well as possible applications of DCC. Authored by the "Who's Who" of DCC it spans the whole range of topics: catalysts, sensors, polymers, ligands, receptors, concluding with a look at future developments and perspectives. All set to become the standard text in the field, this one-stop reference contains everything organic, catalytic, polymer, physical and biochemists need to know.
Download or read book Synthesis and Screening of Peptide Libraries for Biological Applications written by Thi Ba Trinh and published by . This book was released on 2014 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Combinatorial chemistry is a powerful tool in medicinal chemistry as well as chemical biology. In this work, we have applied combinatorial chemistry toward the analysis of peptide cyclization, specificity profiling of protein kinases and the identification of novel inhibitors against medicinally important protein targets.
