Catalytic Performance of a Class III Old Yellow Enzyme and Its Cysteine Variants.

ABSTRACT: Class III old yellow enzymes (OYEs) contain a conserved cysteine in their active sites. To address the role of this cysteine in OYE-mediated asymmetric synthesis, we have studied the biocatalytic properties of OYERo2a from Rhodococcus opacus 1CP (WT) as well as its engineered variants C25A, C25S and C25G. OYERo2a in its redox resting state (oxidized form) is irreversibly inactivated by N-methylmaleimide. As anticipated, inactivation does not occur with the Cys variants. Steady-state kinetics with this maleimide substrate revealed that C25S and C25G doubled the turnover frequency (k _cat) while showing increased K _M values compared to WT, and that C25A performed more similar to WT. Applying the substrate 2-cyclohexen-1-one, the Cys variants w

SUBMITTER: Scholtissek A 

PROVIDER: S-EPMC6194350 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature