title: Insight into the impact of two structural calcium ions on the properties of Pleurotus eryngii versatile ligninolytic peroxidase.
abstract: Two structural Ca2+(proximal and distal) is known to be important for ligninolytic peroxidases. However, few studies toward impact of residues involved in two Ca2+on properties of ligninolytic peroxidases have been done, especially the proximal one. In this study, mutants of nine residues involved in liganding two Ca2+of Pleurotus eryngii versatile peroxidase (VP) were investigated. Most mutants almost completely lost activities, except the mutants of proximal Ca2+- S170A and V192T. In comparison with WT (wild type), optimal pH values of S170A, S170D, and V192T shifted from pH 3.0 to pH 3.5. The order of thermal and pH stabilities of WT, V192T, S170A, and S170D is similar to that of their specific activities: WT > V192T > S170A > S170D. The CD (circular dichroism) results of WT and several mutants indicated that mutations had some effects on secondary structures. For the first time, it was observed that the thermostability of ligninolytic peroxidases is related with proximal Ca2+too, and the mutant containing distal Ca2+only was obtained. Our results clearly demonstrated that enzymatic activities, pH and thermal stabilities, Ca2+content, and secondary structures of VP have close relationship with the residues involved in two structural Ca2+.
authors.affiliation: Department of Microbiology and Parasitology, China Medical University, No.77 Puhe Road, Shenyang 110122, Liaoning Province, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Department of Microbiology and Parasitology, China Medical University, No.77 Puhe Road, Shenyang 110122, Liaoning Province, China. Electronic address: lyzheng@cmu.edu.cn.
authors.forename: Lanyan
authors.initials: L
authors.lastname: Zheng
authors.affiliation: National Key Laboratory of Biochemical Engineering, National Engineering Research Center for Biotechnology (Beijing), Key Laboratory of Biopharmaceutical Production & Formulation Engineering, PLA, Institute of Process Engineering, Chinese Academy of Sciences, No. 1 North 2nd Street, Beijing 100190, China. Electronic address: jjli@ipe.ac.cn.
authors.forename: Jian-Jun
authors.initials: JJ
authors.lastname: Li
authors.affiliation: National Key Laboratory of Biochemical Engineering, National Engineering Research Center for Biotechnology (Beijing), Key Laboratory of Biopharmaceutical Production & Formulation Engineering, PLA, Institute of Process Engineering, Chinese Academy of Sciences, No. 1 North 2nd Street, Beijing 100190, China. Electronic address: ygdu@ipe.ac.cn.
authors.forename: Yuguang
authors.initials: Y
authors.lastname: Du
mesh_terms: D002118:Calcium / Q000737:chemistry*; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008024:Ligands; D008031:Lignin / Q000737:chemistry; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010544:Peroxidases / Q000737:chemistry*; D020076:Pleurotus / Q000201:enzymology*; D017433:Protein Structure, Secondary; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature
chemical_list: D005656:Fungal Proteins; D008024:Ligands; D008031:Lignin; D010544:Peroxidases; D002118:Calcium
keywords: Secondary structure; Site-directed mutagenesis; Structural calcium ion; Versatile peroxidase; pH and thermal stability