Overview
- Editors:
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Frances H. Arnold
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California Institute of Technology, Pasadena
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George Georgiou
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University of Texas at Austin, Austin
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Table of contents (31 protocols)
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Genetic Selections
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- Jessica L. Sneeden, Lawrence A. Loeb
Pages 3-10
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- Manel Camps, Lawrence A. Loeb
Pages 11-18
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- Ranga N. Venkatesan, Lawrence A. Loeb
Pages 19-26
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- Jijumon Chelliserrykattil, Andrew D. Ellington
Pages 27-43
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- Andreas Martin, Franz X. Schmid, Volker Sieber
Pages 57-70
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- Gary W. Rudgers, Timothy Palzkill
Pages 71-81
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Screens for Enzymes
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- Oriana Salazar, Lianhong Sun
Pages 85-97
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- Thomas Bulter, Volker Sieber, Miguel Alcalde
Pages 99-107
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- Alexander V. Tobias, John M. Joern
Pages 109-115
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- Patrick C. Cirino, Radu Georgescu
Pages 117-125
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- Holger Berk, Robert J. Lebbink
Pages 127-135
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- Christopher R. Otey, John M. Joern
Pages 141-148
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- Anton Glieder, Peter Meinhold
Pages 157-170
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About this book
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.
Reviews
"...covers a considerable number of protocols for a broad range of enzymes...very useful..." - ChemBioChem
Editors and Affiliations
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California Institute of Technology, Pasadena
Frances H. Arnold
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University of Texas at Austin, Austin
George Georgiou
