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Protein Evolution In The Presence Of An Unnatural Amino Acid
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Book Synopsis Protein Evolution in the Presence of an Unnatural Amino Acid by : Amrita Singh
Download or read book Protein Evolution in the Presence of an Unnatural Amino Acid written by Amrita Singh and published by . This book was released on 2012 with total page 430 pages. Available in PDF, EPUB and Kindle. Book excerpt: The field of protein engineering has been greatly augmented by the expansion of the genetic code using unnatural amino acids as well as the development of cell-free synthesis systems with high protein yield. Cell-free synthesis systems have improved considerably since they were first described almost 40 years ago. Residue specific incorporation of non-canonical amino acids into proteins is usually performed in vivo using amino acid auxotrophic strains and replacing the natural amino acid with an unnatural amino acid analog. Herein, we present an amino acid depleted cell-free protein synthesis system that can be used to study residue specific replacement of a natural amino acid by an unnatural amino acid analog. This system combines high protein expression yields with a high level of analog substitution in the target protein. To demonstrate the productivity and efficacy of a cell-free synthesis system for residue-specific incorporation of unnatural amino acids in vitro, we use this system to show that 5-fluorotryptophan and 6-fluorotryptophan substituted streptavidin retain the ability to bind biotin despite protein wide replacement of a natural amino acid for the amino acid analog. We envisage this amino acid-depleted cell-free synthesis system being an economical and convenient format for the high-throughput screening of a myriad of amino acid analogs with a variety of protein targets for the study and functional characterization of proteins substituted with unnatural amino acids when compared to the currently employed in vivo format. We use this amino acid depleted cell-free synthesis system for the directed evolution of streptavidin, a protein that finds wide application in molecular biology and biotechnology. We evolve streptavidin using in vitro compartmentalization in emulsions to bind to desthiobiotin and find, at the conclusion of our experiment, that our evolved streptavidin variants are capable of binding to both biotin and desthiobiotin equally well. We also discover a set of mutations for streptavidin that are potentially powerful stabilizing mutations that we believe will be of great use to the greater research community.
Download or read book Non-Natural Amino Acids written by and published by Academic Press. This book was released on 2009-07-24 with total page 334 pages. Available in PDF, EPUB and Kindle. Book excerpt: By combining the tools of organic chemistry with those of physical biochemistry and cell biology, Non-Natural Amino Acids aims to provide fundamental insights into how proteins work within the context of complex biological systems of biomedical interest. The critically acclaimed laboratory standard for 40 years, Methods in Enzymology is one of the most highly respected publications in the field of biochemistry. Since 1955, each volume has been eagerly awaited, frequently consulted, and praised by researchers and reviewers alike. With more than 400 volumes published, each Methods in Enzymology volume presents material that is relevant in today's labs -- truly an essential publication for researchers in all fields of life sciences. Demonstrates how the tools and principles of chemistry combined with the molecules and processes of living cells can be combined to create molecules with new properties and functions found neither in nature nor in the test tube Presents new insights into the molecular mechanisms of complex biological and chemical systems that can be gained by studying the structure and function of non-natural molecules Provides a "one-stop shop" for tried and tested essential techniques, eliminating the need to wade through untested or unreliable methods
Download or read book ספר ילקוט יהושע written by and published by . This book was released on 1933 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis Micro and Nanofabrication Using Self-Assembled Biological Nanostructures by : Jaime Castillo-León
Download or read book Micro and Nanofabrication Using Self-Assembled Biological Nanostructures written by Jaime Castillo-León and published by William Andrew. This book was released on 2014-09-10 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Self-assembled nanostructures based on peptides and proteins have been investigated and presented as biomaterials with an impressive potential for a broad range of applications such as microfabrication, biosensing platforms, drug delivery systems, bioelectronics and tissue reparation. Through self-assembly peptides can give rise to a range of well-defined nanostructures such as nanotubes, nanofibers, nanoparticles, nanotapes, gels and nanorods. However, there are challenges when trying to integrate these biological nanostructures in the development of sensing devices or drug-delivery systems - challenges such as controlling the size during synthesis, the stability in liquid environments and manipulation. In "Micro and Nanofabrication Using Self-assembled Biological Nanostructures" the options and challenges when using self-assembled peptide nanostructures in micro and nanofabrication are discussed. The publication covers different ways to manipulate, deposit and immobilize on specific locations these biological nanostructures in order to use them in the fabrication of new structures or as part of biosensing platforms. Examples where researchers used biological nanostructures for those types of applications are provided. Finally, future applications are discussed as well as parameters to accelerate and expand the use of these biological building blocks in nano- and micro-fabrication processes by taking advantage of their impressive properties such as low-cost and short synthesis time.
Book Synopsis Engineering the Genetic Code by : Nediljko Budisa
Download or read book Engineering the Genetic Code written by Nediljko Budisa and published by John Wiley & Sons. This book was released on 2006-05-12 with total page 312 pages. Available in PDF, EPUB and Kindle. Book excerpt: The ability to introduce non-canonical amino acids in vivo has greatly expanded the repertoire of accessible proteins for basic research and biotechnological application. Here, the different methods and strategies to incorporate new or modified amino acids are explained in detail, including a lot of practical advice for first-time users of this powerful technique. Novel applications in protein biochemistry, genomics, biotechnology and biomedicine made possible by the expansion of the genetic code are discussed and numerous examples are given. Essential reading for all molecular life scientists who want to stay ahead in their research.
Book Synopsis Site-specific Incorporation of Unnatural Amino Acid Into Protein in Vivo by : Kiket Avi Omer Makoubi
Download or read book Site-specific Incorporation of Unnatural Amino Acid Into Protein in Vivo written by Kiket Avi Omer Makoubi and published by . This book was released on 2012 with total page 506 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis Protein Engineering by : Caroline Koehrer
Download or read book Protein Engineering written by Caroline Koehrer and published by Springer Science & Business Media. This book was released on 2009-01-07 with total page 351 pages. Available in PDF, EPUB and Kindle. Book excerpt: Site-specific mutagenesis of DNA, developed some thirty years ago, has proven to be one of the most important advances in biology. By allowing the site-specific replacement of any amino acid in a protein with one of the other nineteen amino acids, it ushered in the new era of "Protein Engineering". The field of protein engineering has, however, evolved rapidly since then and the last fifteen years have witnessed remarkable advances through the use of new chemical, biochemical and molecular biological tools towards the synthesis and manipulation of proteins. The chapters included in this book reflect the rapid evolution of protein engineering and its many applications in basic research, biotechnology, material sciences and therapy. This book will provide the reader with an introduction to state-of the-art concepts and methods and will be of use to anyone interested in the study of proteins, in academia as well as in industry.
Book Synopsis Unnatural Amino Acid Incorporation for Genetic Code Expansion in Mammalian Cells by : Jeffrey Kunio Takimoto
Download or read book Unnatural Amino Acid Incorporation for Genetic Code Expansion in Mammalian Cells written by Jeffrey Kunio Takimoto and published by . This book was released on 2011 with total page 150 pages. Available in PDF, EPUB and Kindle. Book excerpt: The genetic code of most organisms was evolved to encode 20 amino acids. Although the ability to encode 20 amino acids provides the basis to translate proteins necessary for life, researchers are also limited to these 20 amino acids for conventional site-directed mutagenesis. The ability to encode unnatural amino acids provides researchers the ability to circumvent the limitation imposed by the genetic code. Genetically encoding unnatural amino acids provides researchers the means to not only mimic naturally occurring posttranslational modifications but also the ability to encode amino acids with new physical or chemical properties to study biological processes. The incorporation of unnatural amino acids into proteins had been developed in Escherichia coli and also in yeast. We have developed a methodology to genetically incorporate unnatural amino acids in mammalian cells in response to an amber codon (UAG). The incorporation of unnatural amino acids is high in E. coli and yeast, but the incorporation in mammalian cells is relatively low. In addition to developing the system to incorporate unnatural amino acids in mammalian cells, we have also improved suppression efficiencies by modifying the synthetase and unnatural amino acid. To incorporate unnatural amino acids in response to an amber codon, the tRNA anticodon is mutated from a GUA to a CUA. We were able to show that engineering the anticodon-binding domain of the synthetase could enhance the recognition of the tRNA and thus increased suppression efficiencies. Furthermore, by masking the carboxyl group of the amino acid by an ester group, we were able to increase the bioavailability of an unnatural amino acid to further increase suppression efficiencies. Most evolved synthetases aminoacylate unnatural amino acids that are structurally similar to the native substrate of the wild-type synthetase. We were able to adapt Methanosarcina mazei pyrrolysine synthetase (PylRS) to charge a considerable disparate amino acid from its native substrate, O-methyl-L-tyrosine. In addition, the X-ray crystal structure was solved for the evolved PylRS complexed with O-methyl-L-tyrosine at 1.75Å. This multifaceted approach provides the basis to engineer the PylRS to incorporate a significantly diverse selection of unnatural amino acids than previously anticipated.
Book Synopsis The Aminoacyl-tRNA Synthetases by : Michael Ibba
Download or read book The Aminoacyl-tRNA Synthetases written by Michael Ibba and published by CRC Press. This book was released on 2005-04-01 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: By virtue of their role as catalysts of the aminoacylation reaction, the aminoacyl-tRNA synthetases ensure that the first step of translation is performed quickly and accurately. In this volume of 36 separate chapters, the many facets of this ancient and ubiquitous family are reviewed, including their surprising structural diversity, enzymology, tRNA interaction properties, and curious alternative functions. These chapters illustrate the degree to which the aminoacyl-tRNA synthetases employ a variety of mechanisms to carry out both the standard functions related to the synthesis of aminoacylated tRNA for protein synthesis, as well as the surprising functions associated with amino acid biosynthesis, cytokine function, and even the processivity of DNA replication. Other chapters explore the regulation of their synthesis, their role in disease, and their prospects as targets for antibacterial therapeutics. This monograph will be a valuable resource for all scientists interested in the fundamentals of protein synthesis from both a basic research and clinical perspective, as well as the relation of translational components to the evolution of the genetic code.
Book Synopsis Directed Enzyme Evolution by : Frances H. Arnold
Download or read book Directed Enzyme Evolution written by Frances H. Arnold and published by Springer Science & Business Media. This book was released on 2008-02-02 with total page 381 pages. Available in PDF, EPUB and Kindle. Book excerpt: Directed evolution comprises two distinct steps that are typically applied in an iterative fashion: (1) generating molecular diversity and (2) finding among the ensemble of mutant sequences those proteins that perform the desired fu- tion according to the specified criteria. In many ways, the second step is the most challenging. No matter how cleverly designed or diverse the starting library, without an effective screening strategy the ability to isolate useful clones is severely diminished. The best screens are (1) high throughput, to increase the likelihood that useful clones will be found; (2) sufficiently sen- tive (i. e. , good signal to noise) to allow the isolation of lower activity clones early in evolution; (3) sufficiently reproducible to allow one to find small improvements; (4) robust, which means that the signal afforded by active clones is not dependent on difficult-to-control environmental variables; and, most importantly, (5) sensitive to the desired function. Regarding this last point, almost anyone who has attempted a directed evolution experiment has learned firsthand the truth of the dictum “you get what you screen for. ” The protocols in Directed Enzyme Evolution describe a series of detailed p- cedures of proven utility for directed evolution purposes. The volume begins with several selection strategies for enzyme evolution and continues with assay methods that can be used to screen enzyme libraries. Genetic selections offer the advantage that functional proteins can be isolated from very large libraries s- ply by growing a population of cells under selective conditions.
Book Synopsis Site-Specific Protein Labeling by : Arnaud Gautier
Download or read book Site-Specific Protein Labeling written by Arnaud Gautier and published by Humana Press. This book was released on 2015-01-06 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: This detailed volume provides in-depth protocols for protein labeling techniques and applications, with an additional focus on general background information on the design and generation of the organic molecules used for the labeling step. Chapters provide protocols for labeling techniques and applications, with an additional focus on general background information on the design and generation of the organic molecules used for the labeling step. 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. Authoritative and practical, Site-Specific Protein Labeling: Methods and Protocols provides a comprehensive overview on the most relevant and established labeling methodologies, and helps researchers to choose the most appropriate labeling method for their biological question.
Book Synopsis Combinatorial Library by : Lisa B. English
Download or read book Combinatorial Library written by Lisa B. English and published by Springer Science & Business Media. This book was released on 2008-02-04 with total page 380 pages. Available in PDF, EPUB and Kindle. Book excerpt: The continued successes of large- and small-scale genome sequencing projects are increasing the number of genomic targets available for drug d- covery at an exponential rate. In addition, a better understanding of molecular mechanisms—such as apoptosis, signal transduction, telomere control of ch- mosomes, cytoskeletal development, modulation of stress-related proteins, and cell surface display of antigens by the major histocompatibility complex m- ecules—has improved the probability of identifying the most promising genomic targets to counteract disease. As a result, developing and optimizing lead candidates for these targets and rapidly moving them into clinical trials is now a critical juncture in pharmaceutical research. Recent advances in com- natorial library synthesis, purification, and analysis techniques are not only increasing the numbers of compounds that can be tested against each specific genomic target, but are also speeding and improving the overall processes of lead discovery and optimization. There are two main approaches to combinatorial library production: p- allel chemical synthesis and split-and-mix chemical synthesis. These approaches can utilize solid- or solution-based synthetic methods, alone or in combination, although the majority of combinatorial library synthesis is still done on solid support. In a parallel synthesis, all the products are assembled separately in their own reaction vessels or microtiter plates. The array of rows and columns enables researchers to organize the building blocks to be c- bined, and provides an easy way to identify compounds in a particular well.
Book Synopsis Translation Mechanisms by : Jacques Lapointe
Download or read book Translation Mechanisms written by Jacques Lapointe and published by Springer Science & Business Media. This book was released on 2003-07-31 with total page 476 pages. Available in PDF, EPUB and Kindle. Book excerpt: Translation Mechanisms provides investigators and graduate students with overviews of recent developments in the field of protein biosynthesis that are fuelled by the explosive and synergic growth of structural biology, genomics, and bioinformatics. The outstanding progress in our understanding of the structure, dynamics, and evolution of the prokaryotic and eukaryotic translation machinery, as well as applications in medicine and biotechnology, are described in 26 chapters covering recent discoveries on: -the subtleties of tRNA aminoacylation with natural and unnatural amino acids. -the control of mRNA stability, a key step of gene regulation. -ribosome structure and function, in the era of the atomic-crystal resolution of the ribosome. -the regulation of the biosynthesis of the translational machinery components. -the action of a variety of inhibitors of translation and the prospect for clinical studies.
Book Synopsis Chemical Approaches to the Synthesis of Peptides and Proteins by : Paul Lloyd-Williams
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.
Book Synopsis Protein Engineering by : Huimin Zhao
Download or read book Protein Engineering written by Huimin Zhao and published by John Wiley & Sons. This book was released on 2021-08-23 with total page 41 pages. Available in PDF, EPUB and Kindle. Book excerpt: A one-stop reference that reviews protein design strategies to applications in industrial and medical biotechnology Protein Engineering: Tools and Applications is a comprehensive resource that offers a systematic and comprehensive review of the most recent advances in the field, and contains detailed information on the methodologies and strategies behind these approaches. The authors—noted experts on the topic—explore the distinctive advantages and disadvantages of the presented methodologies and strategies in a targeted and focused manner that allows for the adaptation and implementation of the strategies for new applications. The book contains information on the directed evolution, rational design, and semi-rational design of proteins and offers a review of the most recent applications in industrial and medical biotechnology. This important book: Covers technologies and methodologies used in protein engineering Includes the strategies behind the approaches, designed to help with the adaptation and implementation of these strategies for new applications Offers a comprehensive and thorough treatment of protein engineering from primary strategies to applications in industrial and medical biotechnology Presents cutting edge advances in the continuously evolving field of protein engineering Written for students and professionals of bioengineering, biotechnology, biochemistry, Protein Engineering: Tools and Applications offers an essential resource to the design strategies in protein engineering and reviews recent applications.
Book Synopsis Molecular Switches by : Ben L. Feringa
Download or read book Molecular Switches written by Ben L. Feringa and published by John Wiley & Sons. This book was released on 2011-08-04 with total page 814 pages. Available in PDF, EPUB and Kindle. Book excerpt: Täglich benutzen wir Schalter, um strombetriebene Geräte an- und abzuschalten und kein Compuer würde ohne sie funktionieren. Nach den gleichen Prinzipien funktionieren auch molekulare Schalter, die unter dem Einfluß ihrer Umwelt zwischen zwei definierten Zuständen wechseln können. Im Gegensatz zu den gewöhnlichen Schaltern sind molekulare Schalter aber außerordentlich klein und ihre Anwendung in der Nanotechnologie, Biomedizin und im Computerchipdesign öffnet neue Horizonte. Im vorliegenden Zweibänder berichten Herausgeber und Autoren über molekulare Schalter aus Katenanen und Rotaxanen, Fulgiden, Flüssigkristallen und Polypeptiden. Die Bandbreite der behandelten Themen reicht von chiroptischen Schaltern über multifunktionale Systeme bis hin zu molekularen logischen Schaltungen. Chemiker und Materialwissenschaftler in Industrie und Hochschule, die sich für einen der innovativsten Bereiche ihrer Wissenschaft interessieren, werden dieses Buch mit Gewinn lesen!
Book Synopsis Exploring and Expanding the Protein Universe with Non-Canonical Amino Acids by : Gustavo Fuertes
Download or read book Exploring and Expanding the Protein Universe with Non-Canonical Amino Acids written by Gustavo Fuertes and published by Frontiers Media SA. This book was released on 2023-11-01 with total page 123 pages. Available in PDF, EPUB and Kindle. Book excerpt: The site-specific incorporation of unnatural or non-canonical amino acids (ncAAs) into proteins is a universally important tool for systems bioengineering at the interface of chemistry, biology, and biotechnology. The synergistic use of ncAA and related technologies (e.g. Xeno nucleic acids) should enable: i) New opportunities to manipulate, design and elucidate protein structure, dynamics, and function. ii) A deeper understanding of natural and evolved translational systems and their importance for artificial biology. iii) The synthesis of novel biopolymers, creating a solid basis for synthetic cells, which is also an important technology in the production of new classes of medically relevant protein-based scaffolds. Research on reprogrammed protein translation has now reached an experimental and intellectual maturity: more than 200 ncAA (i.e. more than ten times larger variety than standard amino acids) have been introduced into proteins using different routes: genetic code expansion (GCE), selective pressure incorporation (SPI), chemical mutagenesis, protein semi-synthesis, and peptide synthesis.
