Computational Design of Multisubstrate Enzyme Specificity


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
Antony D. St-Jacques, Marie-Ève C. Eyahpaise, Roberto A. Chica,ACS Catalysis (2019) Computational Design of Multisubstrate Enzyme Specificity
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1IYE 2003-05-06 1.82 CRYSTAL STRUCTURE OF ESCHELICHIA COLI BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE
1I1K 2001-07-04 2.1 CRYSTAL STRUCTURE OF ESCHELICHIA COLI BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE.
1IYD 2003-05-06 2.15 CRYSTAL STRUCTURE OF ESCHELICHIA COLI BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE
1I1L 2001-07-04 2.4 CRYSTAL STRUCTURE OF ESCHELICHIA COLI BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE.
1I1M 2001-07-04 2.4 CRYSTAL STRUCTURE OF ESCHERICHIA COLI BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE.
1A3G 1998-05-27 2.5 BRANCHED-CHAIN AMINO ACID AMINOTRANSFERASE FROM ESCHERICHIA COLI

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
300.0 A,B,C Branched-chain-amino-acid aminotransferase P0AB82 ILVE_ECO57
300.0 A,B,C Branched-chain-amino-acid aminotransferase P0AB81 ILVE_ECOL6
300.0 A,B,C Branched-chain-amino-acid aminotransferase P0AB80 ILVE_ECOLI
294.29999999999995 A,B,C Branched-chain-amino-acid aminotransferase P0A1A6 ILVE_SALTI
294.29999999999995 A,B,C Branched-chain-amino-acid aminotransferase P0A1A5 ILVE_SALTY