Structure-based rational design to enhance the solubility and thermostability of a bacterial laccase Lac15

PLoS One. 2014 Jul 18;9(7):e102423. doi: 10.1371/journal.pone.0102423. eCollection 2014.

Abstract

Bacterial laccases are ideal alternatives of fungal laccases for specific industrial applications due to specific characteristics such as alkalescence dependence and high chloride tolerance. However, some bacterial laccases presented as inclusion bodies when expressing in Escherichia coli and showed thermal instability. In this study, rational design was employed to enhance the solubility and the thermostablity of the bacterial laccase Lac15-His6 based on the crystal structure obtained previously. After deletion of His-tag and residues323-332, the obtained Lac15D was completely expressed in soluble form even at a higher temperature of 28°C, compared to only 50% of Lac15-His6 expressed solubly at 16°C. It showed a two-time higher activity at temperatures lower than 35°C and a half-life increasing from 72 min to 150 min at 45°C. When used in chromogenic reactions, Lac15D showed constant activity toward dye precursors and their combinations under alkaline conditions, demonstrating its application potential in hair coloring biotechnology.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / chemistry*
  • Catalysis
  • Chromogenic Compounds / metabolism
  • Crystallography, X-Ray
  • Escherichia coli
  • Hair Dyes / metabolism
  • Hot Temperature
  • Hydrogen-Ion Concentration
  • Laccase / chemistry*
  • Models, Molecular
  • Protein Conformation
  • Protein Engineering
  • Protein Stability
  • Recombinant Fusion Proteins / chemistry
  • Solubility
  • Structure-Activity Relationship
  • Substrate Specificity

Substances

  • Bacterial Proteins
  • Chromogenic Compounds
  • Hair Dyes
  • Recombinant Fusion Proteins
  • Laccase

Grants and funding

This work was funded by the National High Technology Research and Development Program of China (2011AA09070305 to YX); the National Natural Science Foundation of China (31370114 to YX); the Scientific Research Foundation for Returned Scholars, the Ministry of Education of China (to YX); the Natural Science Foundation of Anhui Province (1208085QC63 to JY); the Introduction Program of Academic and Technology Leaders (32030066 to ZF); and the Innovative Research Team Program of 211 Project in Anhui University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.