Reversible control of enantioselectivity by the length of ketone substituent in biocatalytic reduction

Afifa Ayu Koesoema, Yosuke Sugiyama, Kotchakorn T. Sriwong, Zichang Xu, Samantha Verina, Daron M. Standley, Miki Senda, Toshiya Senda, Tomoko Matsuda

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

Enzyme engineering has been widely employed to tailor the substrate specificity and enantioselectivity of enzymes. In this study, we mutated Trp288, an unconserved residue in the small binding pocket of an acetophenone reductase from Geotrichum candidum NBRC 4597 (GcAPRD). Trp288 mutants showed substrate specificity expansion towards bulky-bulky ketones and enantioselectivity alteration which was highly dependent on the substrate substituent length. In aliphatic ketone reduction, enantioselectivity inverted from (S) to (R) when one of the substituents to the carbonyl carbon was elongated from propyl to butyl or pentyl. The best (R)-selective mutant, Trp288Val, achieved the reduction of 3-heptanone to its corresponding (R)-alcohol with 97% ee. Our docking simulation suggested that when enantioselectivity inverted to (R), only pro-R binding poses were productive. Gly94 played an important role to stabilize the butyl or pentyl group for their productive pro-R poses. Interestingly, when the substituent was further elongated, the enantioselectivity inverted back to the (S) form.

Original languageEnglish
Pages (from-to)9529-9541
Number of pages13
JournalApplied Microbiology and Biotechnology
Volume103
Issue number23-24
DOIs
Publication statusPublished - Dec 1 2019
Externally publishedYes

Keywords

  • Alcohol dehydrogenase
  • Asymmetric reduction
  • Docking simulation
  • Enantioselectivity inversion
  • Enzyme engineering

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

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