Directed Evolution of a Cp*Rh^III -Linked Biohybrid Catalyst Based on a Screening Platform with Affinity Purification.

ABSTRACT: Directed evolution of Cp*Rh^III -linked nitrobindin (NB), a biohybrid catalyst, was performed based on an in vitro screening approach. A key aspect of this effort was the establishment of a high-throughput screening (HTS) platform that involves an affinity purification step employing a starch-agarose resin for a maltose binding protein (MBP) tag. The HTS platform enables efficient preparation of the purified MBP-tagged biohybrid catalysts in a 96-well format and eliminates background influence of the host E. coli cells. Three rounds of directed evolution and screening of more than 4000 clones yielded a Cp*Rh^III -linked NB(T98H/L100K/K127E) variant with a 4.9-fold enhanced activity for the cycloaddition of acetophenone oximes with alkynes. It is confirmed that this HTS

SUBMITTER: Kato S 

PROVIDER: S-EPMC7894531 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature