title: Effect of protein structure on laccase-catalyzed protein oligomerization.
abstract: Laccase-catalyzed oligomerization of proteins was studied using Trametes hirsuta laccase (ThL) and coactosin as a model system. The reaction mechanism was elucidated using free amino acids and the tripeptide Gly-Leu-Tyr as substrates. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and high-performance liquid chromatography (HPLC) as well as oxygen consumption measurements and SDS-PAGE were used to study the reactions. Of the 15 selected amino acids, ThL was found to oxidize tryptophan (Trp), tyrosine (Tyr), and cysteine (Cys), of which the reactions with Tyr and Cys have been described earlier. ThL was able to link four full-length coactosins, whereas coactosin that was truncated from its C-terminus remained unpolymerized. Of the four tyrosine residues present in coactosin, only the tyrosine in the C-terminus was found to be reactive. Polymerization between tyrosine side-chains was unambiguously shown using different oligomers of Gly-Leu-Tyr as parent ions in MALDI-TOF/TOF MS fragment ion analyses.
authors.affiliation: VTT, P.O. Box 1000, FIN-02044 VTT, Finland. Maija.Mattinen@vtt.fi
authors.forename: Maija-Liisa
authors.initials: ML
authors.lastname: Mattinen
authors.forename: Maarit
authors.initials: M
authors.lastname: Hellman
authors.forename: Perttu
authors.initials: P
authors.lastname: Permi
authors.forename: Karin
authors.initials: K
authors.lastname: Autio
authors.forename: Nisse
authors.initials: N
authors.lastname: Kalkkinen
authors.forename: Johanna
authors.initials: J
authors.lastname: Buchert
mesh_terms: D001487:Basidiomycota / Q000201:enzymology*; D002384:Catalysis; D004591:Electrophoresis, Polyacrylamide Gel; D042845:Laccase / Q000737:chemistry* / Q000378:metabolism; D008840:Microfilament Proteins / Q000737:chemistry; D008958:Models, Molecular; D009842:Oligopeptides / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000737:chemistry
chemical_list: C485551:Cotl1 protein, mouse; D008840:Microfilament Proteins; D009842:Oligopeptides; D014443:Tyrosine; C104293:glycyl-leucyl-tyrosine; D042845:Laccase