Three-dimensional structure of an alkaline xylanase Xyn11A-LC from alkalophilic Bacillus sp. SN5 and improvement of its thermal performance by introducing arginines substitutions

Biotechnol Lett. 2014 Jul;36(7):1495-501. doi: 10.1007/s10529-014-1512-7. Epub 2014 Mar 30.

Abstract

The alkaline xylanase Xyn11A-LC from the alkalophilic Bacillus sp. SN5 was expressed in E. coli, purified and crystallized. The crystal structure was determined at a resolution of 1.49 Å. Xyn11A-LC has the β-jelly roll structure typical of family 11 xylanases. To improve its thermostability and thermophilicity, a mutant SB3 was constructed by introducing three arginines on the different sides of the protein surface. SB3 increased the optimum temperature by 5 °C. The wild type and SB3 had the half-lives of 22 and 68 min at 65 °C at pH 8.0 (Tris/HCl buffer), respectively. CD spectroscopy revealed that the melting temperature (T m) of the wild type and SB3 were 55.3 and 66.9 °C, respectively. These results showed that the introduction of arginines enhance the thermophilicity and thermostability of Xyn11A-LC.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution*
  • Bacillus / enzymology*
  • Bacillus / genetics
  • Circular Dichroism
  • Crystallography, X-Ray
  • Enzyme Stability
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Hydrogen-Ion Concentration
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Spectrum Analysis
  • Temperature
  • Transition Temperature
  • Xylosidases / chemistry*
  • Xylosidases / genetics
  • Xylosidases / metabolism*

Substances

  • Recombinant Proteins
  • Xylosidases