Directed enzyme evolution : screening and selection methods / edited by Frances H. Arnold and George Georgiou.

Contributor(s): Arnold, Frances Hamilton | Georgiou, George
Material type: TextTextSeries: Methods in molecular biology (Clifton, N.J.): v. 230.Publisher: Totowa, N.J. : Humana Press, ©2003Description: 1 online resource (xv, 383 pages) : illustrations (some color)Content type: text Media type: computer Carrier type: online resourceISBN: 1592593968; 9781592593965Subject(s): Molecular evolution -- Laboratory manuals | Gene libraries -- Laboratory manuals | Protein engineering -- Laboratory manuals | Genetic engineering -- Laboratory manuals | Directed Molecular Evolution -- methods | Enzymes | Evolution, Molecular | Gene Library | Protein Engineering -- methods | Enzymes -- Manuels de laboratoire | Technologie des protéines -- Manuels de laboratoire | Évolution moléculaire -- Manuels de laboratoire | Banques de gènes -- Manuels de laboratoire | Manipulations génétiques -- Manuels de laboratoire | SCIENCE -- Biotechnology | Gene libraries | Genetic engineering | Molecular evolution | Protein engineering | Protein engineering | DNA | Enzymen | evolutie | evolution | enzymen | enzymes | moleculaire biologie | molecular biology | directed evolution | Molecular Biology (General) | Moleculaire biologie (algemeen)Genre/Form: Electronic books. | Laboratory manuals. Additional physical formats: Print version:: Directed enzyme evolution.DDC classification: 660.6/34 LOC classification: QP601 | .D47 2003ebOnline resources: Click here to access online
Contents:
pt. 1. Genetic selections -- Genetic complementation protocols / Jessica L. Sneeden -- Use of Pol 1-Deficient E. coli for functional complementation of DNA polymerase / Manel Camps -- Selection of novel eukaryotic DNA polymerases by mutagenesis and genetic complementation of yeast / Ranga N. Venkatesan -- Autogene selections / jijumon Chelliserrykattil -- Selection for soluble proteins via fusion with chloramphenicol acetyltransferase / Volker Sieber -- Proside: a phage-based method for selecting thermostable proteins / Andreas Martin -- Minimization of proteins by random fragmentation and selection / Gary W. Rudgers -- pt. 2. Screens for enzymes -- Evaluating a screen and analysis of mutant libraries / Oriana Salazar -- Screening mutant libraries in Saccharomyces cerevisiae / Thomas Bulter -- Solid-phase screening using digital image analysis / Alexander V. Tobias -- Screening for thermostability / Patrick C. Cirino -- High-throughput screening of mutant a-amylase libraries for increased activity at 129C / Holger Berk -- High-throughput carbon monoxide binding assay for cytochromes P450 / Christopher R. Otey -- High-throughput screen for aromatic hydroxylation / Christopher R. Otey -- Colorimetric screen for aliphatic hydroxylation by cytochrome P450 using p-ntirophenyl-substituted alkanes / Edgardo T. Farinas -- High-throughput screens based on NAD(P)H depletion / Anton Glieder -- High-throughput tetramethylbenzidine (TMB) screen for peroxidases / Radu Georgescu -- Screen for oxidases by detection of hydrogen peroxide with horseradish peroxidase / Lianhong Sun -- Colorimetric dehydrogenase screen based on NAD(P)H generation / Kimberly M. Mayer -- Colorimetric assays for screening laccases / Miguel Alcalde -- pH sensing agar plate assays for esterolytic enzyme activity / Karl E. Griswold -- A pH-indicator-based screen for hydrolytic haloalkane dehalogenase / Huimin Zhao -- Detection of aromatic a-hydroxyketones with tetrazolium salts / Michael Breuer -- Selection of heat-stable Clostridium cellulovorans cellulases after in vivo recombination / Koichiro Murashima -- Screening and selection strategies for disulfide isomerase activity /Ronald Lafond -- An overview of high-throughput screening systems for enantioselective enyzmatic transformations / Manfred T. Reetz -- Select protocols of high-throughput ee-screening systems for assaying enantioselective enzymes / Manfred T. Reetz -- Directed evolution of the substrate specifications of a site-specific recombinase and an aminoacyl-tRNA synthetase using fluorescence-activated cell sorting (FACS) / Stephen W. Santoro -- Calmodulin-tagged phage and two-filter sandwich assays for the identification of enzymatic activities /Christian Heinis -- High-throughput FACS method for directed evolution of substrate specificity / Mark J. Olsen -- Improving protein folding efficiency by directed evolution using GFP folding reporter / Geoffrey S. Waldo.
Summary: Directed evolution, the application of evolutionary design to enzyme engineering, requires effective screening strategies to isolate those proteins that perform a desired function from the libraries generated by the techniques. In Directed Enzyme Evolution: Screening and Selection Methods, seasoned practitioners from many leading laboratories describe their leading and readily reproducible screening strategies for isolating useful clones. These techniques have been optimized for sensitivity, high throughput, and robustness, and are of proven utility for directed evolution purposes. The assays presented use a variety of techniques, including genetic complementation, microtiter plates, solid-phase screens with colorimetric substrates, and flow cytometric screens. There are also representative examples of how phage libraries may be interrogated for enzymatic activity. Each protocol contains detailed step-by-step instructions and many notes on how best to deal with the problems that may occur. An accompanying volume, Directed Evolution Library Creation: Methods and Protocols (ISBN 1-58829-285-1), describes readily reproducible methods for the creation of mutated DNA molecules and DNA libraries. Taken together, Directed Enzyme Evolution: Screening and Selection Methods and Directed Evolution Library Creation: Methods and Protocols capture for newcomers and more experienced investigators alike all the key methods for using directed protein evolution to better understand protein structure-function relationships, to discover new enzymes and therapeutic proteins, and to design new assays suitable for specific applications.
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Includes bibliographical references and index.

pt. 1. Genetic selections -- Genetic complementation protocols / Jessica L. Sneeden -- Use of Pol 1-Deficient E. coli for functional complementation of DNA polymerase / Manel Camps -- Selection of novel eukaryotic DNA polymerases by mutagenesis and genetic complementation of yeast / Ranga N. Venkatesan -- Autogene selections / jijumon Chelliserrykattil -- Selection for soluble proteins via fusion with chloramphenicol acetyltransferase / Volker Sieber -- Proside: a phage-based method for selecting thermostable proteins / Andreas Martin -- Minimization of proteins by random fragmentation and selection / Gary W. Rudgers -- pt. 2. Screens for enzymes -- Evaluating a screen and analysis of mutant libraries / Oriana Salazar -- Screening mutant libraries in Saccharomyces cerevisiae / Thomas Bulter -- Solid-phase screening using digital image analysis / Alexander V. Tobias -- Screening for thermostability / Patrick C. Cirino -- High-throughput screening of mutant a-amylase libraries for increased activity at 129C / Holger Berk -- High-throughput carbon monoxide binding assay for cytochromes P450 / Christopher R. Otey -- High-throughput screen for aromatic hydroxylation / Christopher R. Otey -- Colorimetric screen for aliphatic hydroxylation by cytochrome P450 using p-ntirophenyl-substituted alkanes / Edgardo T. Farinas -- High-throughput screens based on NAD(P)H depletion / Anton Glieder -- High-throughput tetramethylbenzidine (TMB) screen for peroxidases / Radu Georgescu -- Screen for oxidases by detection of hydrogen peroxide with horseradish peroxidase / Lianhong Sun -- Colorimetric dehydrogenase screen based on NAD(P)H generation / Kimberly M. Mayer -- Colorimetric assays for screening laccases / Miguel Alcalde -- pH sensing agar plate assays for esterolytic enzyme activity / Karl E. Griswold -- A pH-indicator-based screen for hydrolytic haloalkane dehalogenase / Huimin Zhao -- Detection of aromatic a-hydroxyketones with tetrazolium salts / Michael Breuer -- Selection of heat-stable Clostridium cellulovorans cellulases after in vivo recombination / Koichiro Murashima -- Screening and selection strategies for disulfide isomerase activity /Ronald Lafond -- An overview of high-throughput screening systems for enantioselective enyzmatic transformations / Manfred T. Reetz -- Select protocols of high-throughput ee-screening systems for assaying enantioselective enzymes / Manfred T. Reetz -- Directed evolution of the substrate specifications of a site-specific recombinase and an aminoacyl-tRNA synthetase using fluorescence-activated cell sorting (FACS) / Stephen W. Santoro -- Calmodulin-tagged phage and two-filter sandwich assays for the identification of enzymatic activities /Christian Heinis -- High-throughput FACS method for directed evolution of substrate specificity / Mark J. Olsen -- Improving protein folding efficiency by directed evolution using GFP folding reporter / Geoffrey S. Waldo.

Print version record.

Directed evolution, the application of evolutionary design to enzyme engineering, requires effective screening strategies to isolate those proteins that perform a desired function from the libraries generated by the techniques. In Directed Enzyme Evolution: Screening and Selection Methods, seasoned practitioners from many leading laboratories describe their leading and readily reproducible screening strategies for isolating useful clones. These techniques have been optimized for sensitivity, high throughput, and robustness, and are of proven utility for directed evolution purposes. The assays presented use a variety of techniques, including genetic complementation, microtiter plates, solid-phase screens with colorimetric substrates, and flow cytometric screens. There are also representative examples of how phage libraries may be interrogated for enzymatic activity. Each protocol contains detailed step-by-step instructions and many notes on how best to deal with the problems that may occur. An accompanying volume, Directed Evolution Library Creation: Methods and Protocols (ISBN 1-58829-285-1), describes readily reproducible methods for the creation of mutated DNA molecules and DNA libraries. Taken together, Directed Enzyme Evolution: Screening and Selection Methods and Directed Evolution Library Creation: Methods and Protocols capture for newcomers and more experienced investigators alike all the key methods for using directed protein evolution to better understand protein structure-function relationships, to discover new enzymes and therapeutic proteins, and to design new assays suitable for specific applications.

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