Characterization of two novel thermostable esterases from Thermoanaerobacterium thermosaccharolyticum

Wenqian Li; Hao Shi; Huaihai Ding; Liangliang Wang; Yu Zhang; Xun Li; Fei Wang

doi:10.1016/j.pep.2018.04.010

Characterization of two novel thermostable esterases from Thermoanaerobacterium thermosaccharolyticum

Protein Expr Purif. 2018 Dec:152:64-70. doi: 10.1016/j.pep.2018.04.010. Epub 2018 Apr 22.

Authors

Wenqian Li¹, Hao Shi², Huaihai Ding³, Liangliang Wang⁴, Yu Zhang⁵, Xun Li⁶, Fei Wang⁷

Affiliations

¹ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China; College of Life Science and Food Engineering, Huaiyin Institute of Technology, Huaian 223003, China. Electronic address: liwenqian@126.com.
² Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China; College of Life Science and Food Engineering, Huaiyin Institute of Technology, Huaian 223003, China. Electronic address: ilyshihao@163.com.
³ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China. Electronic address: dinghuaihai@qq.com.
⁴ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China. Electronic address: wangliangliang32316@126.com.
⁵ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China. Electronic address: yuzhang@njfu.edu.cn.
⁶ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China. Electronic address: xunlee@163.com.
⁷ Jiangsu Co-Innovation Center of Efficient Processing and Utilization of Forest Resources, Nanjing Forestry University, Nanjing 210037, China; College of Chemical Engineering, Jiangsu Key Lab of Biomass-Based Green Fuels and Chemicals, Nanjing Forestry University, Nanjing 210037, China. Electronic address: hgwf01@njfu.edu.cn.

PMID: 29684442
DOI: 10.1016/j.pep.2018.04.010

Abstract

This paper first describes characterization of two thermostable esterases (ThLip1 and ThLip2) from the thermophilic bacterium Thermoanaerobacterium thermosaccharolyticum DSM 571. The recombinant esterase ThLip1 was active at 80 °C, pH 6.5 and maintained approx. 85% of original activity after 2 h incubation at 75 °C. Kinetic parameters, K_m, V_max and k_cat/K_m for 4-Nitrophenyl caprylate (pNPC) were 3.52 ± 0.47 mM, 191.18 ± 1.82 μmol min^-1 mg^-1 and 20.80 ± 0.07 mM^-1 s^-1, respectively. The purified recombinant esterase ThLip2 was optimally active at pH 6.5 and 75 °C and it was stable against a pH range of 6.0-8.0 possessing 2 h half-life at 80 °C. Kinetic experiments at 75 °C with pNPC as a substrate gave a K_m of 3.37 mM, V_max of 578.14 μmol min^-1 mg^-1and k_cat of 231.2 s^-1. The hydrolysis of linalyl acetate were carried out using ThLip1 and ThLip2 as catalyst, affording linalool yields over 140 mg/l in 10 h.

Keywords: Esterase; Hydrolysis; Linalyl acetate; Thermoanaerobacterium thermosaccharolyticum.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acyclic Monoterpenes
Bacterial Proteins / genetics
Bacterial Proteins / isolation & purification
Bacterial Proteins / metabolism*
Caprylates / metabolism
Chromatography, Affinity
Cloning, Molecular
Enzyme Assays
Enzyme Stability
Escherichia coli / enzymology
Escherichia coli / genetics*
Esterases / genetics
Esterases / isolation & purification
Esterases / metabolism*
Gene Expression
Genetic Vectors / chemistry
Genetic Vectors / metabolism
Hot Temperature
Hydrogen-Ion Concentration
Hydrolysis
Isoenzymes / genetics
Isoenzymes / isolation & purification
Isoenzymes / metabolism
Kinetics
Monoterpenes / metabolism*
Recombinant Proteins / genetics
Recombinant Proteins / isolation & purification
Recombinant Proteins / metabolism
Substrate Specificity
Thermoanaerobacterium / chemistry
Thermoanaerobacterium / enzymology*

Substances

Acyclic Monoterpenes
Bacterial Proteins
Caprylates
Isoenzymes
Monoterpenes
Recombinant Proteins
4-nitrophenyloctanoate
linalyl acetate
linalool
Esterases