title: Structural diversity in the dandelion (Taraxacum officinale) polyphenol oxidase family results in different responses to model substrates.
abstract: Polyphenol oxidases (PPOs) are ubiquitous type-3 copper enzymes that catalyze the oxygen-dependent conversion of o-diphenols to the corresponding quinones. In most plants, PPOs are present as multiple isoenzymes that probably serve distinct functions, although the precise relationship between sequence, structure and function has not been addressed in detail. We therefore compared the characteristics and activities of recombinant dandelion PPOs to gain insight into the structure-function relationships within the plant PPO family. Phylogenetic analysis resolved the 11 isoenzymes of dandelion into two evolutionary groups. More detailed in silico and in vitro analyses of four representative PPOs covering both phylogenetic groups were performed. Molecular modeling and docking predicted differences in enzyme-substrate interactions, providing a structure-based explanation for grouping. One amino acid side chain positioned at the entrance to the active site (position HB2+1) potentially acts as a "selector" for substrate binding. In vitro activity measurements with the recombinant, purified enzymes also revealed group-specific differences in kinetic parameters when the selected PPOs were presented with five model substrates. The combination of our enzyme kinetic measurements and the in silico docking studies therefore indicate that the physiological functions of individual PPOs might be defined by their specific interactions with different natural substrates.
authors.affiliation: Department of Plant Biology and Biotechnology, Westphalian Wilhelms-University of Münster, Münster, Germany.
authors.forename: Mareike E
authors.initials: ME
authors.lastname: Dirks-Hofmeister
authors.affiliation: Department of Plant Biology and Biotechnology, Westphalian Wilhelms-University of Münster, Münster, Germany.
authors.forename: Ratna
authors.initials: R
authors.lastname: Singh
authors.affiliation: Department of Plant Biology and Biotechnology, Westphalian Wilhelms-University of Münster, Münster, Germany.
authors.forename: Christine M
authors.initials: CM
authors.lastname: Leufken
authors.affiliation: Department of Chemistry and Physics, Indiana State University, Terre Haute, Indiana, United States of America.
authors.forename: Jennifer K
authors.initials: JK
authors.lastname: Inlow
authors.affiliation: Department of Plant Biology and Biotechnology, Westphalian Wilhelms-University of Münster, Münster, Germany.
authors.forename: Bruno M
authors.initials: BM
authors.lastname: Moerschbacher
mesh_terms: D000595:Amino Acid Sequence; D005075:Biological Evolution; D002384:Catalysis; D020134:Catalytic Domain / Q000235:genetics; D004156:Catechol Oxidase / Q000737:chemistry* / Q000235:genetics; D003300:Copper / Q000737:chemistry; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010100:Oxygen; D010802:Phylogeny; D011809:Quinones / Q000737:chemistry; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D031205:Taraxacum / Q000737:chemistry*
chemical_list: D011809:Quinones; D003300:Copper; D004156:Catechol Oxidase; D010100:Oxygen