Journal
NATURE CATALYSIS
Volume 5, Issue 2, Pages 136-+Publisher
NATURE PORTFOLIO
DOI: 10.1038/s41929-022-00743-0
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Funding
- Michigan Economic Development Corporation
- Michigan Technology Tri-Corridor [085P1000817]
- Office of the Vice Chancellor for Research and Graduate Education at the University of Wisconsin-Madison
- Wisconsin Alumni Research Foundation
- National Institute of Health [DP2-GM137417]
- Morgridge Institute for Research-Metabolism Theme Fellowship
- NIH Biotechnology Training Grant [T32-GM008349]
- Bender Fund
- US Department of Energy, Office of Science, Office of Basic Energy Sciences [W-31-109-Eng-38]
- NSF [CHE-1919350]
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Enantioselective C-C bond-forming reactions are underdeveloped in biocatalysis. This study presents an engineered decarboxylative aldolase enzyme that facilitates the synthesis of non-standard gamma-hydroxy amino acids. The enzyme shows improved activity through directed evolution and engineering, and demonstrates high efficiency in whole-cell biocatalysis reactions.
Enantioselective C-C bond-forming reactions are underdeveloped in the biocatalysis toolbox. Now, engineering an efficient and promiscuous decarboxylative aldolase enzyme provides a solution to facilitate the convenient synthesis of non-standard gamma-hydroxy amino acids from simple building blocks. Enzymes are renowned for their catalytic efficiency and selectivity. Despite the wealth of carbon-carbon bond-forming transformations in traditional organic chemistry and nature, relatively few C-C bond-forming enzymes have found their way into the biocatalysis toolbox. Here we show that the enzyme UstD performs a highly selective decarboxylative aldol addition with diverse aldehyde substrates to make non-standard gamma-hydroxy amino acids. We increased the activity of UstD through three rounds of classic directed evolution and an additional round of computationally guided engineering. The enzyme that emerged, UstD(v2.0), is efficient in a whole-cell biocatalysis format. The products are highly desirable, functionally rich bioactive gamma-hydroxy amino acids that we demonstrate can be prepared stereoselectively on the gram scale. The X-ray crystal structure of UstD(v2.0) at 2.25 angstrom reveals the active site and provides a foundation for probing the UstD mechanism.
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