Engineering Enzyme Specificity Using Computational Design of a Defined-Sequence Library
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Lippow-2010-Engineering Enzyme S.pdf
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Author(s)
Lippow, Shaun M.
Moon, Tae Seok
Basu, Subhayu
Yoon, Sang-Hwal
Li, Xiazhen
Chapman, Brad A.
Robison, Keith
Lipovšek, Daša
Prather, Kristala L. Jones
Date Issued
December 2010
Journal
Chemistry and Biology
Publisher
Elsevier
Citation
Lippow, Shaun M., Tae Seok Moon, Subhayu Basu, Sang-Hwal Yoon, Xiazhen Li, Brad A. Chapman, Keith Robison, Daša Lipovšek, and Kristala L.J. Prather. “Engineering Enzyme Specificity Using Computational Design of a Defined-Sequence Library.” Chemistry & Biology 17, no. 12 (December 2010): 1306–1315. © 2010 Elsevier Ltd.
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Final published version
Abstract
Engineered biosynthetic pathways have the potential to produce high-value molecules from inexpensive feedstocks, but a key limitation is engineering enzymes with high activity and specificity for new reactions. Here, we developed a method for combining structure-based computational protein design with library-based enzyme screening, in which inter-residue correlations favored by the design are encoded into a defined-sequence library. We validated this approach by engineering a glucose 6-oxidase enzyme for use in a proposed pathway to convert D-glucose into D-glucaric acid. The most active variant, identified after only one round of diversification and screening of only 10,000 wells, is approximately 400-fold more active on glucose than is the wild-type enzyme. We anticipate that this strategy will be broadly applicable to the discovery of new enzymes for engineered biological pathways.
MIT Department
Massachusetts Institute of Technology. Department of Chemical Engineering
Massachusetts Institute of Technology. Synthetic Biology Center
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Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use.
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DOI of Published Version
https://doi.org/10.1016/j.chembiol.2010.10.012
