title: Heterologous expression and functional characterization of a novel cellulose-disruptive protein LeEXP2 from Lycopersicum esculentum.
abstract: There has been little research reported on the synergism of plant expansins in cellulose hydrolysis because of the difficulty of heterologous expression. In this study, the expansin gene LeExp2 of Lycopersicum esculentum was expressed for the first time in Pichia pastoris to identify its function in vitro. The optimal expression level was obtained by adding PMSF and a commercial protease inhibitor, and the maximum expression level was 70.9 mg/L. The affinity-purified LeEXP2 displayed cellulose-weakening activity and synergism with cellulase, and the reducing sugar yield in the reaction mixture with LeEXP2 was 1.4-9.8-fold that of control. The optimum pH and temperature for reducing-sugar liberation by LeEXP2 and endoglucanase were pH 4.8 and 50 °C. Cu(2+) and Fe(2+) notably increased the synergistic activity of LeEXP2, and Co(2+) enhanced the activity of endoglucanase but failed to improve synergism. LeEXP2 exhibited a high level of robustness to reagents and heat, indicating its potential application for cellulose decomposition. LeEXP2 preferentially bound to cellulose over lignin, and the binding capability to given substrates was not related to the crystallinity. This is the first study to characterize the functional role of LeEXP2 in cellulose degradation.
authors.affiliation: R&D Center for Petrochemical Technology, Tianjin University, Tianjin 300072, China; Department of Biochemical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, People's Republic of China.
authors.forename: Xuewei
authors.initials: X
authors.lastname: Liu
authors.affiliation: R&D Center for Petrochemical Technology, Tianjin University, Tianjin 300072, China.
authors.forename: Cheng
authors.initials: C
authors.lastname: Liu
authors.affiliation: R&D Center for Petrochemical Technology, Tianjin University, Tianjin 300072, China. Electronic address: myy@tju.edu.cn.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Ma
authors.affiliation: R&D Center for Petrochemical Technology, Tianjin University, Tianjin 300072, China.
authors.forename: Jiefang
authors.initials: J
authors.lastname: Hong
authors.affiliation: R&D Center for Petrochemical Technology, Tianjin University, Tianjin 300072, China; State Key Laboratory of Engine, Tianjin University, Tianjin 300072, China.
authors.forename: Minhua
authors.initials: M
authors.lastname: Zhang
mesh_terms: D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D018551:Solanum lycopersicum / Q000235:genetics*; D008958:Models, Molecular; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
chemical_list: D010940:Plant Proteins; D011994:Recombinant Proteins; C095825:expansin protein, plant
keywords: Binding characteristic; Cellulose hydrolysis; Expansin; Heterologous expression; Robustness; Synergistic activity