Biocatalytic Synthesis of Non-Standard Amino Acids by a Decarboxylative Aldol Reaction

07 May 2021, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


The formation of carbon-carbon bonds lies at the heart of organic chemistry, but relatively few C-C bond forming enzymes have found their way into the biocatalysis toolbox. We report that the enzyme UstD performs a highly selective decarboxylative aldol addition with diverse aldehyde substrates to make non-standard, γ-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, UstD2.0, is very efficient in a whole-cell biocatalysis format and readily crystallizes. The X-ray crystal structure of UstD2.0 at 2.25 Å reveals the active site and empowers future studies. The utility of UstD2.0 was demonstrated via the stereoselective gram-scale syntheses of non-standard amino acids.


Non-standard amino acids
Directed evolution
pyridoxal phosphate


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