Functional analysis of a divergent system II protein, Ccs1, involved in c-type cytochrome biogenesis.
The Ccs1 gene, encoding a highly divergent novel component of a system II type c-type cytochrome biogenesis pathway, is encoded by the previously defined CCS1 locus in Chlamydomonas reinhardtii. phoA and lacZalpha bacterial topological reporters were used to deduce a topological model of the Synechocystis sp. 6803 Ccs1 homologue, CcsB. CcsB, and therefore by analogy Ccs1, possesses a large soluble lumenal domain at its C terminus that is tethered in the thylakoid membrane by three closely spaced transmembrane domains in the N-terminal portion of the protein. Molecular analysis of ccs1 alleles reveals that the entire C-terminal soluble domain is essential for Ccs1 function and that a stromal loop appears to be important in vivo, at least for maintenance of Ccs1. Site-directed mutational analysis reveals that a single histidine (His(274)) within the last transmembrane domain, preceding the large lumenal domain, is required for c-type cytochrome assembly, whereas an invariant cysteine residue (Cys(199)) is shown to be non-essential. Ccs1 is proposed to interact with other Ccs components based on its reduced accumulation in ccs2, ccs3, ccs4, and ccsA strains.
Dreyfuss
Beth Welty
BW
Department of Chemistry and Biochemistry, UCLA, Los Angeles, California 90095-1569, USA.
Hamel
Patrice P
PP
Nakamoto
Stacie S
SS
Merchant
Sabeeha
S
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern; D002462:Cell Membrane / Q000378:metabolism; D016825:Chlamydomonas reinhardtii / Q000378:metabolism; D060365:Chloroplast Proteins; D003580:Cytochromes / Q000378:metabolism*; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D006639:Histidine / Q000737:chemistry; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010940:Plant Proteins; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D012313:RNA / Q000378:metabolism; D020524:Thylakoids / Q000378:metabolism; D013997:Time Factors
C109655:Ccs1 protein, Chlamydomonas reinhardtii; D060365:Chloroplast Proteins; D003580:Cytochromes; D010940:Plant Proteins; D015800:Protozoan Proteins; D006639:Histidine; D012313:RNA