Rational selection of circular permutation sites in characteristic regions of the α/β-hydrolase fold enzyme RhEst1

In this study we performed the CP analysis of the typical esterase RhEst1 to explore the CP site-selection strategy of the α/β-hydrolase fold family. A CP library of 97 mutants was generated to identify the effect of CP on three characteristic regions of RhEst1including the flexible cap domain (Region 1), the region around the entrance to substrate binding pocket (Region 2) and the surface-exposed sectors in catalytic domain (Region 3). We found the protein folding, stability and bioactivity of CP variants were altered significantly and the CP sites of active variants were mainly located in the flexible loops. These studies reveal the importance of site-selection for CP and provide more information for CP of other α/β-hydrolases. Graphical abstract
Source: Journal of Molecular Catalysis B: Enzymatic - Category: Biochemistry Source Type: research