Identification of stabilizing point mutations through mutagenesis of destabilized protein libraries.

ABSTRACT: Although there have been recent transformative advances in the area of protein structure prediction, prediction of point mutations that improve protein stability remains challenging. It is possible to construct and screen large mutant libraries for improved activity or ligand binding. However, reliable screens for mutants that improve protein stability do not yet exist, especially for proteins that are well folded and relatively stable. Here, we demonstrate that incorporation of a single, specific, destabilizing mutation termed parent inactivating mutation into each member of a single-site saturation mutagenesis library, followed by screening for suppressors, allows for robust and accurate identification of stabilizing mutations. We carried out fluorescence-activated cell sorting of such a

SUBMITTER: Ahmed S 

PROVIDER: S-EPMC8971944 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature