Investigation of Spectroscopic Intermediates during Copper-Binding and TPQ Formation in Wild-Type and Active-Site Mutants of a Copper-Containing Amine Oxidase from Yeast

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• 作者：Joanne E. Dove ; Benjamin Schwartz ; Neal K. Williams ; and Judith P. Klinman
• 刊名：Biochemistry
• 出版年：2000
• 出版时间：April 4, 2000
• 年：2000
• 卷：39
• 期：13
• 页码：3690 - 3698
• 全文大小：171K
• 年卷期：v.39,no.13(April 4, 2000)
• ISSN：1520-4995

文摘

Copper amine oxidases possess the unusual ability to generate autocatalytically their organiccofactor, which is subsequently utilized in turnover. This cofactor, 2,4,5-trihydroxyphenylalanine quinone(TPQ), is formed within the active site of these enzymes by the oxidation of a single tyrosine residue. Invitro, copper(II) and oxygen are both necessary and sufficient for the conversion of tyrosine to TPQ. Inthis study, the biogenesis of TPQ has been characterized in an amine oxidase from Hansenula polymorphaexpressed as the apo-enzyme in Escherichia coli. With the WT enzyme, optical absorbances which arecopper or oxygen dependent are observed and characterized. Active-site mutants are used to investigatefurther the nature of these spectral species. Evidence is presented which suggests that tyrosine is activatedfor reaction with oxygen by liganding to Cu(II). In the following paper in this issue [Schwartz, B., Dove,J. E., and Klinman, J. P. (2000) Biochemistry 39, 3699-3707], the initial reaction of precursor proteinwith oxygen is characterized kinetically. Taken together, the available data suggest a mechanism for theoxidation of tyrosine to TPQ where the role of the copper is to activate substrate.