Structural Snapshots from the Oxidative Half-reaction of a Copper Amine Oxidase IMPLICATIONS FOR O

• Published in: The Journal of biological chemistry Vol. 288; no. 39; pp. 28409 - 28417
• Main Authors: Johnson, Bryan J., Yukl, Erik T., Klema, Valerie J., Klinman, Judith P., Wilmot, Carrie M.
• Format: Journal Article
• Language: English
• Published: 9650 Rockville Pike, Bethesda, MD 20814, U.S.A American Society for Biochemistry and Molecular Biology 12.08.2013
• Subjects: Enzymology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.501791

Background: Copper amine oxidases activate O 2 either at the copper center or aminoquinol cofactor. Results: Catalytic intermediates from the oxidative half-reaction are structurally and spectroscopically characterized. Conclusion: The mechanism of O 2 activation may depend on accessibility dictated by two conformers of the quinone cofactor. Significance: Structural changes that inform on catalytic mechanism have been revealed in the ubiquitous copper amine oxidases. The mechanism of molecular oxygen activation is the subject of controversy in the copper amine oxidase family. At their active sites, copper amine oxidases contain both a mononuclear copper ion and a protein-derived quinone cofactor. Proposals have been made for the activation of molecular oxygen via both a Cu(II)-aminoquinol catalytic intermediate and a Cu(I)-semiquinone intermediate. Using protein crystallographic freeze-trapping methods under low oxygen conditions combined with single-crystal microspectrophotometry, we have determined structures corresponding to the iminoquinone and semiquinone forms of the enzyme. Methylamine reduction at acidic or neutral pH has revealed protonated and deprotonated forms of the iminoquinone that are accompanied by a bound oxygen species that is likely hydrogen peroxide. However, methylamine reduction at pH 8.5 has revealed a copper-ligated cofactor proposed to be the semiquinone form. A copper-ligated orientation, be it the sole identity of the semiquinone or not, blocks the oxygen-binding site, suggesting that accessibility of Cu(I) may be the basis of partitioning O 2 activation between the aminoquinol and Cu(I).