Unknown

Dataset Information

0

Optimizing glycosyltransferase specificity via "hot spot" saturation mutagenesis presents a catalyst for novobiocin glycorandomization.


ABSTRACT: A comprehensive two-phase "hot spot" saturation mutagenesis strategy for the rapid evolution of glycosyltransferase (GT) specificity for nonnatural acceptors is described. Specifically, the application of a high-throughput screen (based on the fluorescent acceptor umbelliferone) was used to identify key amino acid hot spots that contribute to GT proficiency and/or promiscuity. Saturation mutagenesis of the corresponding hot spots facilitated the utilization of a lower-throughput screen to provide OleD prodigy capable of efficiently glycosylating the nonnatural acceptor novobiocic acid with an array of unique sugars. Incredibly, even in the absence of a high-throughput screen for novobiocic acid glycosylation, this approach rapidly led to improvements in the desired catalytic activity of several hundred-fold.

SUBMITTER: Williams GJ 

PROVIDER: S-EPMC2813856 | biostudies-literature | 2008 Apr

REPOSITORIES: biostudies-literature

altmetric image

Publications

Optimizing glycosyltransferase specificity via "hot spot" saturation mutagenesis presents a catalyst for novobiocin glycorandomization.

Williams Gavin J GJ   Goff Randal D RD   Zhang Changsheng C   Thorson Jon S JS  

Chemistry & biology 20080401 4


A comprehensive two-phase "hot spot" saturation mutagenesis strategy for the rapid evolution of glycosyltransferase (GT) specificity for nonnatural acceptors is described. Specifically, the application of a high-throughput screen (based on the fluorescent acceptor umbelliferone) was used to identify key amino acid hot spots that contribute to GT proficiency and/or promiscuity. Saturation mutagenesis of the corresponding hot spots facilitated the utilization of a lower-throughput screen to provid  ...[more]

Similar Datasets

| S-EPMC8745346 | biostudies-literature
| S-EPMC3837798 | biostudies-literature
| S-EPMC4978272 | biostudies-literature
| S-EPMC4398508 | biostudies-literature
| S-EPMC4959485 | biostudies-literature
2019-02-15 | GSE126550 | GEO
| S-EPMC5666567 | biostudies-literature
2017-11-27 | GSE105785 | GEO
2024-04-27 | GSE265942 | GEO
| S-EPMC3562065 | biostudies-literature