Improvement of Selenomonas ruminantium β-xylosidase thermal stability by replacing buried free cysteines via site directed mutagenesis.

Improvement of Selenomonas ruminantium β-xylosidase thermal stability by replacing buried free cysteines via site directed mutagenesis.
Int J Biol Macromol. 2019 Jun 17;:
Authors: Dehnavi E, Moeini S, Akbarzadeh A, Dabirmanesh B, Siadat SOR, Khajeh K
Abstract
β-xylosidase is an essential enzyme for breakdown of xylan to d-xylose. It has a significant potential application value for medicine, food, paper and pulp, and biofuel industries. Due to the negative consequences caused by buried free cysteine residues, mutational substitution of such residues is often accompanied by a notable increase in thermal stability. To characterize the role of cysteine residues in the structure, function and stability of Selenomonas ruminantium β-d-Xylosidase (SXA), we prepared and evaluated wild-type and four cysteines- deficient SXA proteins. Buried cysteine residues were replaced with. In comparison with the wild-type, the Km values of the mutants remained relatively constant while their kcat values decreased. The C101V and C286V displayed higher thermal stability than the wild-type at 55 and 60 °C. Conformational changes of the secondary and tertiary structure as derived from circular dichroism and fluorescence spectroscopy revealed that changing a buried cysteine to a hydrophobic residue could lead to an increase in thermal stability with minimal perturbation of the wild-type protein structure. In addition to experimental methods, the stabili...

Source: International Journal of Biological Macromolecules - June 16, 2019 Category: Biochemistry Authors: Dehnavi E, Moeini S, Akbarzadeh A, Dabirmanesh B, Siadat SOR, Khajeh K Tags: Int J Biol Macromol Source Type: research