Abstract

The engineering of multi-substrate enzyme specificity is highly desirable to foster the application of biocatalysis in industry. Here, we develop a multistate computational protein design methodology called multi-chemical state analysis (MCSA) that can optimize enzyme sequences on large structural ensembles for productive binding of multiple target substrates. Using MCSA, we redesigned E. coli branched-chain amino acid aminotransferase to accept both α-ketoglutarate and the non-native substrate L-histidine. Screening of a designed combinatorial library comprising 32 mutants for enhanced L-histidine transamination activity yielded four variants displaying up to ≈200-fold improvements to kcat/KM. MCSA opens the door to the design of broad-specificity biocatalysts and multi-substrate enzymes displaying tailored specificity.

Submission Details

ID: DddCjJY34

Submitter: Antony St-Jacques

Submission Date: May 7, 2020, 7:42 a.m.

Version: 1

Publication Details

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