Biochemical and Spectroscopic Characterization of the Covalent Binding of Heme to Cytochrome b6

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• 作者：Catherine de Vitry ; Alain Desbois ; Virginie Redeker ; Francesca Zito ; and Francis-Andr&eacute ; Wollman
• 刊名：Biochemistry
• 出版年：2004
• 出版时间：April 6, 2004
• 年：2004
• 卷：43
• 期：13
• 页码：3956 - 3968
• 全文大小：276K
• 年卷期：v.43,no.13(April 6, 2004)
• ISSN：1520-4995

文摘

The three-dimensional structure of the cytochrome b₆f complex disclosed the unexpectedpresence of a new heme c_i [Stroebel, D., Choquet, Y., Popot, J.-L., and Picot, D. (2003) Nature 426,413-418; Kurisu, G., Zhang, H., Smith, J. L., and Cramer, W. A. (2003) Science 302, 1009-1014].Here we present a biochemical, spectroscopic, and mutagenesis study of this unusual heme binding inChlamydomonas reinhardtii. As predicted by the structure data, we identify a Cys³⁵-containing proteolyticfragment (Tyr²⁵-Lys¹¹¹) from cytochrome b₆ as a peptide that covalently binds a heme. Resonance Ramanspectra of cyt b₆f complexes show particular frequencies in s/gifchars/nu.gif" BORDER=0 >₂, s/gifchars/nu.gif" BORDER=0 >₃, s/gifchars/nu.gif" BORDER=0 >₄, and s/gifchars/nu.gif" BORDER=0 >₈ regions that identify thisextra heme as a ferrous c'-like heme under a five-coordinated high-spin state. The set of frequencies isconsistent with a coordination by either a water molecule or a hydroxide ion. Other changes in resonanceRaman bands, observed in the mid- and low-frequency regions, point to a modification in conformationand/or environment of at least one b heme methyl and/or propionate group. Site-directed mutagenesis ofapocytochrome b₆, leading to a Cys³⁵Val substitution, generates Chlamydomonas strains that are unableto assemble cytochrome b₆f complexes. On the basis of the mutant phenotype, we discuss the participation,in the covalent binding of heme c_i, of the nuclear CCB factors that we identified previously as controllingthe apo to holo conversion of cytochrome b₆ [Kuras, R., de Vitry, C., Choquet, Y., Girard-Bascou, J.,Culler, D., Büschlen, S., Merchant, S., and Wollman, F.-A. (1997) J. Biol. Chem. 272, 32427-32435].