Directed Evolution Library Creation
Directed Evolution Library Creation

Author: Frances H. Arnold

Publisher: Humana Press

Published: 2010-11-10

Total Pages: 0

ISBN-13: 9781617374715

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Biological systems are very special substrates for engineering—uniquely the products of evolution, they are easily redesigned by similar approaches. A simple algorithm of iterative cycles of diversification and selection, evolution works at all scales, from single molecules to whole ecosystems. In the little more than a decade since the first reported applications of evolutionary design to enzyme engineering, directed evolution has matured to the point where it now represents the centerpiece of industrial biocatalyst development and is being practiced by thousands of academic and industrial scientists in com- nies and universities around the world. The appeal of directed evolution is easy to understand: it is conceptually straightforward, it can be practiced without any special instrumentation and, most important, it frequently yields useful solutions, many of which are totally unanticipated. Directed evolution has r- dered protein engineering readily accessible to a broad audience of scientists and engineers who wish to tailor a myriad of protein properties, including th- mal and solvent stability, enzyme selectivity, specific activity, protease s- ceptibility, allosteric control of protein function, ligand binding, transcriptional activation, and solubility. Furthermore, the range of applications has expanded to the engineering of more complex functions such as those performed by m- tiple proteins acting in concert (in biosynthetic pathways) or as part of mac- molecular complexes and biological networks.

Directed Enzyme Evolution
Directed Enzyme Evolution

Author: Frances H. Arnold

Publisher: Springer Science & Business Media

Published: 2008-02-02

Total Pages: 381

ISBN-13: 1592593968

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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.

Directed Enzyme Evolution: Advances and Applications
Directed Enzyme Evolution: Advances and Applications

Author: Miguel Alcalde

Publisher: Springer

Published: 2017-02-14

Total Pages: 286

ISBN-13: 3319504134

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This book focuses on some of the most significant advances in enzyme engineering that have been achieved through directed evolution and hybrid approaches. On the 25th anniversary of the discovery of directed evolution, this volume is a tribute to the pioneers of this thrilling research field, and at the same time provides a comprehensive overview of current research and the state of the art. Directed molecular evolution has become the most reliable and robust method to tailor enzymes, metabolic pathways or even whole microorganisms with improved traits. By mirroring the Darwinian algorithm of natural selection on a laboratory scale, new biomolecules of invaluable biotechnological interest can now be engineered in a manner that surpasses the boundaries of nature. The volume is divided into two sections, the first of which provides an update on recent successful cases of enzyme ensembles from different areas of the biotechnological spectrum, including tryptophan synthases, unspecific peroxygenases, phytases, therapeutic enzymes, stereoselective enzymes and CO2-fixing enzymes. This section also provides information on the directed evolution of whole cells. The second section of the book summarizes a variety of the most applicable methods for library creation, together with the future trends aimed at bringing together directed evolution and in silico/computational enzyme design and ancestral resurrection.

Protein Engineering
Protein Engineering

Author: Huimin Zhao

Publisher: John Wiley & Sons

Published: 2021-08-23

Total Pages: 41

ISBN-13: 3527344705

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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.

Directed Evolution Library Creation
Directed Evolution Library Creation

Author: Elizabeth M.J. Gillam

Publisher: Springer

Published: 2016-09-17

Total Pages: 0

ISBN-13: 9781493953295

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Directed Evolution Library Creation: Methods and Protocols, Second Edition presents user-friendly protocols for both proven strategies and cutting-edge approaches for the creation of mutant gene libraries for directed evolution. As well as experimental methods, information on current computational approaches is provided in a user-friendly format that will allow researchers to make informed choices without needing to comprehend the full technical details of each algorithm. Directed evolution has become a fundamental approach for engineering proteins to enhance activity and explore structure-function relationships, and has supported the rapid development of the field of synthetic biology over the last decade. Divided into three convenient sections, topics include point mutagenesis strategies, recombinatorial methods wherein genetic diversity is sourced from multiple parental genes that are combined via either homology-dependent or -independent techniques and a variety of computational methods to guide the design and analysis of mutant libraries. Written in the 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 protocols and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Directed Evolution Library Creation: Methods and Protocols, Second Edition will serve as a reliable manual for both novice and experienced protein engineers and synthetic biologists and will enable further technical innovation and the exploitation of directed evolution for a deeper understanding of protein design and function.

Directed Evolution
Directed Evolution

Author: Andrew Currin

Publisher: Springer Nature

Published: 2022-06-21

Total Pages: 281

ISBN-13: 1071621521

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This volume explores the latest techniques used by researchers to study directed evolution (DE) at each stage of the Design-Build-Test-Learn cycle. Chapters in this book cover topics such as designing overlap extension PCR primers for protein mutagenesis; antha-guided automation of Darwin assembly for the construction of bespoke gene libraries; rapid cloning of random mutagenesis libraries using PTO-Quickstep; and DE of glycosyltransferases by a single-cell screening method. 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, Directed Evolution: Methods and Protocols is a valuable resource for scientists and researchers who are interested in learning more about this field and incorporating these studies into new experimental workflows.

Biodefense in the Age of Synthetic Biology
Biodefense in the Age of Synthetic Biology

Author: National Academies of Sciences, Engineering, and Medicine

Publisher: National Academies Press

Published: 2019-01-05

Total Pages: 189

ISBN-13: 0309465184

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Scientific advances over the past several decades have accelerated the ability to engineer existing organisms and to potentially create novel ones not found in nature. Synthetic biology, which collectively refers to concepts, approaches, and tools that enable the modification or creation of biological organisms, is being pursued overwhelmingly for beneficial purposes ranging from reducing the burden of disease to improving agricultural yields to remediating pollution. Although the contributions synthetic biology can make in these and other areas hold great promise, it is also possible to imagine malicious uses that could threaten U.S. citizens and military personnel. Making informed decisions about how to address such concerns requires a realistic assessment of the capabilities that could be misused. Biodefense in the Age of Synthetic Biology explores and envisions potential misuses of synthetic biology. This report develops a framework to guide an assessment of the security concerns related to advances in synthetic biology, assesses the levels of concern warranted for such advances, and identifies options that could help mitigate those concerns.

Molecular Biomethods Handbook
Molecular Biomethods Handbook

Author: John M. Walker

Publisher: Springer Science & Business Media

Published: 2008-11-04

Total Pages: 1105

ISBN-13: 1603273751

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Recent advances in the biosciences have led to a range of powerful new technologies, particularly nucleic acid, protein and cell-based methodologies. The most recent insights have come to affect how scientists investigate and define cellular processes at the molecular level. This book expands upon the techniques included in the first edition, providing theory, outlines of practical procedures, and applications for a range of techniques. Written by a well-established panel of research scientists, the book provides an up-to-date collection of methods used regularly in the authors’ own research programs.

In the Light of Evolution
In the Light of Evolution

Author: National Academy of Sciences

Publisher:

Published: 2007

Total Pages: 388

ISBN-13:

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The Arthur M. Sackler Colloquia of the National Academy of Sciences address scientific topics of broad and current interest, cutting across the boundaries of traditional disciplines. Each year, four or five such colloquia are scheduled, typically two days in length and international in scope. Colloquia are organized by a member of the Academy, often with the assistance of an organizing committee, and feature presentations by leading scientists in the field and discussions with a hundred or more researchers with an interest in the topic. Colloquia presentations are recorded and posted on the National Academy of Sciences Sackler colloquia website and published on CD-ROM. These Colloquia are made possible by a generous gift from Mrs. Jill Sackler, in memory of her husband, Arthur M. Sackler.