Improvement thermostability of Pseudoalteromonas carrageenovora arylsulfatase by rational design

Yanbing Zhu; Chaochao Qiao; Hebin Li; Lijun Li; Anfeng Xiao; Hui Ni; Zedong Jiang

doi:10.1016/j.ijbiomac.2017.11.014

Improvement thermostability of Pseudoalteromonas carrageenovora arylsulfatase by rational design

Int J Biol Macromol. 2018 Mar:108:953-959. doi: 10.1016/j.ijbiomac.2017.11.014. Epub 2017 Nov 4.

Authors

Yanbing Zhu¹, Chaochao Qiao², Hebin Li³, Lijun Li¹, Anfeng Xiao¹, Hui Ni¹, Zedong Jiang⁴

Affiliations

¹ College of Food and Biological Engineering, Jimei University, Xiamen 361021, China; Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China; Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China; Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China.
² College of Food and Biological Engineering, Jimei University, Xiamen 361021, China.
³ Xiamen Medical College, Xiamen 361008, China.
⁴ College of Food and Biological Engineering, Jimei University, Xiamen 361021, China; Fujian Provincial Key Laboratory of Food Microbiology and Enzyme Engineering, Xiamen 361021, China; Research Center of Food Biotechnology of Xiamen City, Xiamen 361021, China; Key Laboratory of Systemic Utilization and In-depth Processing of Economic Seaweed, Xiamen Southern Ocean Technology Center of China, Xiamen 361021, China. Electronic address: zdjiang@jmu.edu.cn.

PMID: 29113885
DOI: 10.1016/j.ijbiomac.2017.11.014

Abstract

This study aimed to improve the thermostability of arylsulfatase from Pseudoalteromonas carrageenovora. A total of 10 single-site mutants were chosen using the PoPMuSiC program, and two mutants of K253N and P314T showed enhanced thermal stability. By saturation mutagenesis and thermostability analysis, K253H and P314T were the best mutants at the two sites. Combinational mutations of K253H, P314T and H260L were subsequently introduced, and the best mutant of K253H/H260L was selected. Thermal inactivation analysis showed the half-life (t_1/2) value at 55°C for K253H/H260L was 7.7-fold that of the wild-type enzyme (WT), meanwhile this mutant maintained the specific enzyme activity. Structure modeling demonstrated that the additional hydrogen bonds, optimization of surface charge-charge interactions, and increasing of hydrophobic interaction could account for the improved thermostability imparted by K253H/H260L.

Keywords: Arylsulfatase; Pseudoalteromonas carrageenovora; Rational design; Thermostability.

MeSH terms

Amino Acid Substitution
Arylsulfatases / chemistry*
Arylsulfatases / genetics
Arylsulfatases / isolation & purification
Arylsulfatases / metabolism*
Circular Dichroism
Enzyme Activation
Enzyme Stability
Gene Expression
Hydrogen-Ion Concentration
Hydrophobic and Hydrophilic Interactions
Kinetics
Models, Molecular
Mutagenesis, Site-Directed
Mutation
Protein Conformation
Protein Engineering*
Pseudoalteromonas / enzymology*
Pseudoalteromonas / genetics
Temperature

Substances

Arylsulfatases