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In Vivo Incorporation Of Unnatural Amino Acids Into Proteins
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Book Synopsis In Vivo Incorporation of Unnatural Amino Acids Into Proteins by :
Download or read book In Vivo Incorporation of Unnatural Amino Acids Into Proteins written by and published by . This book was released on 2002 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: A method for the site-specific incorporation of unnatural amino acids into proteins in vivo would significantly facilitate studies of the cellular function of proteins, as well as make possible the biosynthesis of unnatural polymers and proteins with novel structures and activities. Our approach consists of the generation of amber suppressor tRNA/aminoacyl-tRNA synthetase pair that are not catalytically competent with all the endogenous Escherichia coli tRNAs an aminoacyl-tRNA synthetases, followed by directed evolution of such orthogonal aminoacyl-tRNA synthetases to alter their amino acid specificities. A new orthogonal suppressor tRNA/aminoacyl-tRNA synthetase pair in E. coli has been derived from the Saccharomyces cerevisiae tRNA (sub Asp) and aspartyl-tRNA syathetase, and the in vitro and in vivo characteristics of this pair were determined. In order to achieve a high specificity for the amino acid, a direct selection for site-specific incorporation of unnatural amino acids into a reporter epitope displayed on the surface of M13 phage has been developed and characterized. Under simulated selection conditions, phage particles displaying aspartate were enriched over 300-fold from a pool of phage displaying asparagine using monoclonal antibodies raised against the aspartate-containing epitope. The direct phage selection offers very high specificity for the amino acid of interest, Which cannot be achieved by conventional methods.
Book Synopsis A General Strategy for Site-specific Incorporation of Unnatural Amino Acids Into Proteins by : Spencer Jay Anthony-Cahill
Download or read book A General Strategy for Site-specific Incorporation of Unnatural Amino Acids Into Proteins written by Spencer Jay Anthony-Cahill and published by . This book was released on 1990 with total page 676 pages. Available in PDF, EPUB and Kindle. Book excerpt:
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
Book Synopsis Unnatural Amino Acids by : Loredano Pollegioni
Download or read book Unnatural Amino Acids written by Loredano Pollegioni and published by Humana Press. This book was released on 2011-10-05 with total page 409 pages. Available in PDF, EPUB and Kindle. Book excerpt: Even though they are present in nature, non-proteinogenic amino acids are usually defined as unnatural or non-natural. Beside their structural diversity, interest in these compounds is due to their occurrence in nature, their biological properties, the analytical aspects, their use as probes, and their incorporation into peptides and proteins, among other reasons. Divided into five convenient sections, Unnatural Amino Acids: Methods and Protocols deals with enzymatic methods used to produce non-natural amino acids, aspects concerning the presence of unnatural amino acids in peptides with antimicrobial properties, genetic incorporation of unnatural amino acids into proteins (yeast and mammalian cells), and detection and quantification of D-amino acids and related enzymes. Written in the highly successful Methods in Molecular BiologyTM series format, chapters contain introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and accessible, Unnatural Amino Acids: Methods and Protocols serves as an ideal guide for scientists and contributes to directing the attention of researchers to the many fields of growing scientific interest in non-natural amino acids.
Book Synopsis Genetic Incorporation of Unnatural Amino Acids Into Proteins by : Isaac Nnamdi Ugwumba
Download or read book Genetic Incorporation of Unnatural Amino Acids Into Proteins written by Isaac Nnamdi Ugwumba and published by . This book was released on 2010 with total page 308 pages. Available in PDF, EPUB and Kindle. Book excerpt: The introduction of unnatural amino acids with novel properties into proteins is potentially a useful tool for understanding and manipulating their functions. The development of an in vitro method for the incorporation of a fluorescent amino acid L-(7-hydroxycoumarin-4-yl)ethylglycine (Hco), into proteins is described here.
Book Synopsis Ex Vivo Incorporation of Unnatural Amino Acids Into Recombinant Proteins by : Sviatlana Smolskaya
Download or read book Ex Vivo Incorporation of Unnatural Amino Acids Into Recombinant Proteins written by Sviatlana Smolskaya and published by . This book was released on 2015 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis Enzyme Engineering by : James Samuelson
Download or read book Enzyme Engineering written by James Samuelson and published by Humana Press. This book was released on 2016-05-01 with total page 266 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book provides guidance to those wishing to create enzyme variants. It covers such topics as a simple method for generating site-specific mutations within bacterial chromosomes and the engineering of two difference types of rare-cutting endonucleases.
Book Synopsis The Development of Intrinsically Fluorescent Unnatural Amino Acids for in Vivo Incorporation Into Proteins by : Itthipol Sungwienwong
Download or read book The Development of Intrinsically Fluorescent Unnatural Amino Acids for in Vivo Incorporation Into Proteins written by Itthipol Sungwienwong and published by . This book was released on 2018 with total page 306 pages. Available in PDF, EPUB and Kindle. Book excerpt: The amino acid acridon-2-ylalanine (Acd) can be a valuable probe of protein dynamics either alone or as part of a Förster resonance energy transfer (FRET) or photo-induced electron transfer (eT) probe pair. We have previously reported the genetic incorporation of Acd by an aminoacyl tRNA synthetase (RS). However, this RS, developed from a library of permissive RSs, also incorporates N-phenyl-amino-phenylalanine (Npf), a trace byproduct of one Acd synthetic route. We have performed negative selections in the presence of Npf and analyzed the selectivity of the resulting AcdRSs by in vivo protein expression and detailed kinetic analyses of the purified RSs. We find that selection conferred a ~50-fold increase in selectivity for Acd over Npf, eliminating incorporation of Npf contaminants, and allowing one to use a high yielding Acd synthetic route for improved overall expression of Acd-containing proteins. More generally, our report also provides a cautionary tale on the use of permissive RSs, as well as a strategy for improving selectivity for the target amino acid. In spite of its utility for studying proteins by fluorescence spectroscopy, Acd can potentially be improved by making it longer wavelength or brighter. We reported the synthesis of Acd core derivatives and their photophysical characterization. We also performed ab initio calculations of the absorption and emission spectra of Acd derivatives, which agree well with experimental measurements. The amino acid aminoacridonylalanine (Aad) was synthesized in forms appropriate for genetic incorporation and peptide synthesis. We show that Aad is a superior FRET acceptor to Acd in a peptide cleavage assay, and that Aad can be activated by an aminoacyl tRNA synthetase for genetic incorporation. Together, these results show that we can use computation to design enhanced Acd derivatives which can be used in peptides and proteins. Finally, the Aad synthesis has been improved and it will be further tested in vivo incorporations into proteins, and alkylated Aad core analogs show improved brightness making their use as amino acids promising.
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.
Book Synopsis In Vivo Incorporation of Unnatural Amino Acids by :
Download or read book In Vivo Incorporation of Unnatural Amino Acids written by and published by . This book was released on 2008 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: The invention provides methods and compositions for in vivo incorporation of unnatural amino acids. Also provided are compositions including proteins with unnatural amino acids.
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 : 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 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 Engineering E. Coli Translational Apparatus by : Thomas John Magliery
Download or read book Engineering E. Coli Translational Apparatus written by Thomas John Magliery and published by . This book was released on 2001 with total page 416 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis A General Method for Site-specific Incorporation of Unnatural Amino Acids Into Proteins by : Christopher J. Noren
Download or read book A General Method for Site-specific Incorporation of Unnatural Amino Acids Into Proteins written by Christopher J. Noren and published by . This book was released on 1989 with total page 7 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Book Synopsis Noncanonical Amino Acids by : Edward A. Lemke
Download or read book Noncanonical Amino Acids written by Edward A. Lemke and published by . This book was released on 2018 with total page 411 pages. Available in PDF, EPUB and Kindle. Book excerpt:
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.
