Thermostabilization of VPR, a kinetically stable cold adapted subtilase, via multiple proline substitutions into surface loops.

ABSTRACT: Protein stability is a widely studied topic, there are still aspects however that need addressing. In this paper we examined the effects of multiple proline substitutions into loop regions of the kinetically stable proteinase K-like serine protease VPR, using the thermostable structural homologue AQUI as a template. Four locations for proline substitutions were chosen to imitate the structure of AQUI. Variants were produced and characterized using differential scanning calorimetry (DSC), circular dichroism (CD), steady state fluorescence, acrylamide fluorescence quenching and thermal inactivation experiments. The final product VPR_ΔC_N3P/I5P/N238P/T265P was greatly stabilized which was achieved without any noticeable detrimental effects to the catalytic efficiency of the enzyme.

SUBMITTER: Oskarsson KR 

PROVIDER: S-EPMC6978356 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature