Veratryl alcohol binding sites of lignin peroxidase from Phanerochaete chrysosporium

Toru Johjima, Hiroyuki Wariishi, Hiroo Tanaka

研究成果: ジャーナルへの寄稿記事

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Possible binding sites of lignin peroxidase (LiP) for veratryl alcohol (VA) were investigated by determining the reactivity of three different chemically modified LiPs against VA when acting (i) as a reducing substrate, (ii) as a rescuing reagent for the rapid conversion of LiPIII* back to native LiP, and (iii) as an enzyme-bound redox mediator. The enzyme was chemically modified to alter its surface properties by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) in the presence and the absence of 2-aminoethanesulfonic acid to introduce sulfo and N-acylurea groups instead of carboxylic groups, respectively. LiP was also modified by N-bromosuccinimide (NBS) to yield Trp-modified enzyme. The spectral characteristics and compound I formation rates of modified LiPs were identical to those of unmodified LiP. The activities for VA oxidation by modified LiPs were significantly reduced but with almost unchanged pH dependencies. Several other substrates including phenolic, anionic, and polymeric substrates were also utilized to characterize the activity of the modified enzymes. Kinetic analysis of these reactions strongly suggests that LiP has at least two substrate oxidation sites, one is Trp 171 for VA oxidation and the other is for anionic substrate oxidation. For VA-supported oxidation of ferric cytochrome c (Cc3+), the VA binding site was estimated to be the same as that of VA oxidation. The reactivities of chemically modified LiPs with excess H2O2 were investigated, indicating the formation of compound III* species. Compound III* species of EDC-LiP and unmodified LiP were converted back to ferric enzymes by adding VA. However, there was a reduced recovery of ferric NBS-LiP, suggesting that the VA binding site for VA-derived rapid reversion of compound III* is located at a different site from that for VA oxidation.

元の言語英語
ページ(範囲)49-57
ページ数9
ジャーナルJournal of Molecular Catalysis - B Enzymatic
17
発行部数2
DOI
出版物ステータス出版済み - 4 25 2002

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All Science Journal Classification (ASJC) codes

  • Catalysis
  • Bioengineering
  • Biochemistry
  • Process Chemistry and Technology

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