title: Extension of polyphenolics by CWPO-C peroxidase mutant containing radical-robust surface active site.
abstract: Expressed as insoluble forms in Escherichia coli, native cationic cell wall peroxidase (CWPO-C) from the poplar tree and mutant variants were successfully reactivated via refolding experiments and used to elucidate the previously presumed existence of an electron transfer (ET) pathway in the CWPO-C structure. Their catalytic properties were fully characterized through various analyses including steady-state kinetic, direct oxidation of lignin macromolecules and their respective stabilities during the polymerization reactions. The analysis results proved that the 74th residue on the CWPO-C surface plays an important role in catalyzing the macromolecules via supposed ET mechanism. By comparing the residual activities of wild-type CWPO-C and mutant 74W CWPO-C after 3 min, mutation of tyrosine 74 residue to tryptophan increased the radical resistance of peroxidase up to ten times dramatically while maintaining its capability to oxidize lignin macromolecules. Furthermore, extension of poly(catechin) as well as lignin macromolecules with CWPO-C Y74W mutant clearly showed that this radical-resistant peroxidase mutant can increase the molecular weight of various kinds of polyphenolics by using surface-located active site. The anti-oxidation activity of the synthesized poly(catechin) was confirmed by xanthine oxidase assay. The elucidation of a uniquely catalytic mechanism in CWPO-C may improve the applicability of the peroxidase/H2O2 catalyst to green polymer chemistry.
authors.forename: L T Mai
authors.initials: LT
authors.lastname: Pham
authors.forename: S Jin
authors.initials: SJ
authors.lastname: Kim
authors.forename: U Suk
authors.initials: US
authors.lastname: Ahn
authors.forename: J Weon
authors.initials: JW
authors.lastname: Choi
authors.forename: B Keun
authors.initials: BK
authors.lastname: Song
authors.forename: Y Hwan
authors.initials: YH
authors.lastname: Kim
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D055162:Biocatalysis; D020134:Catalytic Domain; D002392:Catechin / Q000737:chemistry / Q000378:metabolism; D002412:Cations; D002473:Cell Wall / Q000201:enzymology*; D002850:Chromatography, Gel; D019281:Dimerization; D004795:Enzyme Stability; D005609:Free Radicals / Q000378:metabolism*; D007700:Kinetics; D062105:Molecular Docking Simulation; D009154:Mutation / Q000235:genetics; D010084:Oxidation-Reduction; D009195:Peroxidase / Q000378:metabolism*; D010636:Phenols / Q000737:chemistry / Q000378:metabolism; D058105:Polymerization; D059808:Polyphenols / Q000378:metabolism*; D032107:Populus / Q000201:enzymology; D013053:Spectrophotometry; D040681:Structural Homology, Protein; D013816:Thermodynamics
chemical_list: D000975:Antioxidants; D002412:Cations; D005609:Free Radicals; D010636:Phenols; D059808:Polyphenols; D002392:Catechin; C010559:coniferyl alcohol; D009195:Peroxidase