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020 _a9781592593965
024 7 _a10.1385/1592593968
_2doi
040 _aES-MaUEC
_bspa
_cES-MaUEC
245 1 0 _aDirected Enzyme Evolution
_bScreening and Selection Methods
_cedited by Frances H. Arnold, George Georgiou.
250 _a1st edition 2003
264 1 _aTotowa, NJ
_bHumana Press
_c2003
300 _a1 recurso en línea (XV, 383 páginas)
_b
336 _atexto
_btxt
_2rdacontent
337 _aelectrónico
_bc
_2rdamedia
338 _arecurso electrónico
_bcr
_2rdacarrier
347 _aarchivo de texto
_bPDF
490 0 _aMethods in Molecular Biology
_x1940-6029
_v230
505 0 _aGenetic Selections -- Genetic Complementation Protocols -- Use of Pol I-Deficient E. coli for Functional Complementation of DNA Polymerase -- Selection of Novel Eukaryotic DNA Polymerases by Mutagenesis and Genetic Complementation of Yeast -- Autogene Selections -- Selection for Soluble Proteins via Fusion with Chloramphenicol Acetyltransferase -- Proside -- Minimization of Proteins by Random Fragmentation and Selection -- Screens for Enzymes -- Evaluating a Screen and Analysis of Mutant Libraries -- Screening Mutant Libraries in Saccharomyces cerevisiae -- Solid-Phase Screening Using Digital Image Analysis -- Screening for Thermostability -- High-Throughput Screening of Mutant ?-Amylase Libraries for Increased Activity at 129°C -- High-Throughput Carbon Monoxide Binding Assay for Cytochromes P450 -- High-Throughput Screen for Aromatic Hydroxylation -- Colorimetric Screen for Aliphatic Hydroxylation by Cytochrome P450 Using p-Nitrophenyl-Substituted Alkanes -- High-Throughput Screens Based on NAD(P)H Depletion -- High-Throughput Tetramethylbenzidine (TMB) Screen for Peroxidases -- Screen for Oxidases by Detection of Hydrogen Peroxide with Horseradish Peroxidase -- Colorimetric Dehydrogenase Screen Based on NAD(P)H Generation -- Colorimetric Assays for Screening Laccases -- pH Sensing Agar Plate Assays for Esterolytic Enzyme Activity -- A pH-Indicator-Based Screen for Hydrolytic Haloalkane Dehalogenase -- Detection of Aromatic ?-Hydroxyketones with Tetrazolium Salts -- Selection of Heat-Stable Clostridium cellulovorans Cellulases After In Vitro Recombination -- Screening and Selection Strategies for Disulfide Isomerase Activity -- An Overview of High-Throughput Screening Systems for Enantioselective Enzymatic Transformations -- Select Protocols of High-Throughput ee-Screening Systems for Assaying Enantioselective Enzymes -- Directed Evolution of the Substrate Specificities of a Site-Specific Recombinase and an Aminoacyl-tRNA Synthetase Using Fluorescence-Activated Cell Sorting (FACS) -- Calmodulin-Tagged Phage and Two-Filter Sandwich Assays for the Identification of Enzymatic Activities -- High-Throughput FACS Method for Directed Evolution of Substrate Specificity -- Improving Protein Folding Efficiency by Directed Evolution Using the GFP Folding Reporter.
520 _aDirected 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.
700 1 _aArnold, Frances H
_eeditor literario
_4edt
_4http://id.loc.gov/vocabulary/relators/edt
700 1 _aGeorgiou, George
_eeditor literario
_4edt
_4http://id.loc.gov/vocabulary/relators/edt
776 0 8 _iPrinted edition:
_z9781617374722
776 0 8 _iPrinted edition:
_z9781489938107
776 0 8 _iPrinted edition:
_z9781588292865
856 4 0 _uhttps://go.openathens.net/redirector/universidadeuropea.es?url=https://doi.org/10.1385/1592593968
_z(usuarios Universidad Europea de Valencia)
942 _2lcc
_cLE
988 _aSpringer_Protocols_2003
999 _c233817
_d233817