Molecular Genetic Identification of a Pathway for Heme Binding to Cytochrome b 6

Abstract

Heme binding to cytochrome b 6 is resistant, in part, to denaturing conditions that typically destroy the noncovalent interactions between the b hemes and their apoproteins, suggesting that one of two b hemes of holocytochrome b 6 is tightly bound to the polypeptide. We exploited this property to define a pathway for the conversion of apo-to holocytochrome b 6 , and to identify mutants that are blocked at one step of this pathway.

The Light Reactions: A Guide to Recent Acquisitions for the Picture Gallery

“…The new heme is covalently attached through a single thioether linkage. The existence of an extra heme was unexpected, but the fact that it is covalently attached was satisfying because biochemical analysis had suggested the presence of covalent heme in cytochrome b 6 , and biogenesis studies had indicated that heme association depended on a multistep sequential pathway (Kuras et al, 1997). A quinone analog called stigmatellin is known from biochemical work to interact at or near the Q o site, and its presence in the complex with inhibitor added therefore distinguishes the Q o pocket.…”

“…The presence of the new heme, a chlorophyll, and a carotenoid generates some problems with respect to the biogenesis of the complex, not least of which is the molecular identity of the CCB loci responsible for the step-wise, sequential assembly of the hemes into cytochrome b 6 (Kuras et al, 1997). Given the presence of covalent heme in the b subunit of the respiratory b-c complex in Bacillus subtilis and the conservation in the cytochrome c maturation pathway between the Bacillus sp and chloroplasts, one would predict conservation of the CCB mechanism as well (Kranz et al, 1998;Yu and Le Brun, 1998).…”

Regulatory factors for the assembly of thylakoid membrane protein complexes

“…System IV maturation pathways, namely the assembly of co-factors on complex C subunit B (CCB), are responsible for the attachment of haem ci to apocytochrome b 6 on the stromal side of the thylakoid membrane [97]. At least four protein factors, CCB1 -CCB4, are necessary for this maturation pathway [98].…”

Thylakoid FtsH Protease Contributes to Photosystem II and Cytochromeb 6 fRemodeling inChlamydomonas reinhardtiiunder Stress Conditions

“…Chlamydomonas reinhardtii Strains, Growth Conditions, and Genetic Methods C. reinhardtii mutants have been described previously and were as follows: ccb, {petB-C35V}, {petB-H202Q}, {petB-H187G}, {DpetB}, {DpetA}, {Fud34}, {Fud7}, {D1TR}, arg7, and arg2 (Eversole, 1956;Matagne, 1978;de Vitry et al, 1989de Vitry et al, , 2004Kuras and Wollman, 1994;Kuras et al, 1997Kuras et al, , 2007Minai et al, 2006). Cells were grown at 25°C in Tris-acetate-phosphate (TAP) medium or, when indicated, in minimal medium lacking acetate, pH 7.2, under dim light at 6 mE m 22 s 21 and collected during the exponential phase at 2 3 10 6 cells mL 21 .…”

“…The cytochrome b 6 f complex differs from bc 1 by an additional c i heme located at the quinone reduction site (Kurisu et al, 2003;Stroebel et al, 2003). In recent years, we characterized cofactor assembly to complex C subunit B (CCB) factors involved in the covalent binding of c i heme to Cys-35 of cytochrome b 6 in C. reinhardtii (Kuras et al, 1997(Kuras et al, , 2007 and Arabidopsis (Lezhneva et al, 2008). Mutants lacking c i heme cannot grow photoautotrophically because they show too little accumulation of a dysfunctional b 6 f complex (Saint-Marcoux et al, 2009).…”