CtaG is required for formation of active cytochrome c oxidase in Bacillus subtilis

Abstract

The Gram-positive bacterium Bacillus subtilis contains two respiratory oxidases of the haem-copper superfamily: cytochrome aa 3, which is a quinol oxidase, and cytochrome caa 3, which is a cytochrome c oxidase. Cytochrome c oxidase uniquely contains a di-copper centre, CuA. B. subtilis CtaG is a membrane protein encoded by the same gene cluster as that which encodes the subunits of cytochrome c oxidase. The role of B. subtilis CtaG and orthologous proteins present in many other Gram-positive bacteria has remained unexplored. The sequence of CtaG is unrelated to that of CtaG/Cox11p of proteobacteria and eukaryotic cells. This study shows that B. subtilis CtaG is essential for the formation of active cytochrome caa 3 but is not required for assembly of the core subunits I and II with haem in the membrane and it has no role in the synthesis of active cytochrome aa 3. B. subtilis YpmQ, a homologue to Sco1p of eukaryotic cells, is also a membrane-bound cytochrome c oxidase-specific assembly factor. Properties of CtaG- and YpmQ-deficient mutants were compared. Cells lacking YpmQ showed a low cytochrome c oxidase activity and this defect was suppressed by the supplementation of the growth medium with copper ions. It has previously been proposed that YpmQ/Sco1p is involved in synthesis of the CuA centre. The results of this study are consistent with this proposal but the exact role of YpmQ in assembly of cytochrome c oxidase remains to be elucidated.

Copper-responsive gene expression in the methanotroph Methylosinus trichosporium OB3b

“…These gamma-proteobacterial copD homologues are predicted to have 14 transmembrane helices, and the extra 6 helices are part of a C-terminal domain with some homology to the cytochrome c oxidase caa3 assembly factor (CtaG), which is thought to be involved in the correct assembly of the Cu A site of Bacillus subtilis cytochrome c oxidase caa3 (not to be confused with the Cox11 homologue known as CtaG that is found in Rhodobacter sphaeroides and Paracoccus denitrificans ). 56,57 As with the C-terminal cbb3 -like domain, this domain is particularly well represented in methanotrophs and ammonia oxidizers, and interestingly, a variant with 16 helices is also common in Gram positive bacteria (Fig. S5, ESI†).…”

The CopC Family: Structural and Bioinformatic Insights into a Diverse Group of Periplasmic Copper Binding Proteins

“…In support of this notion, at least 15% of copD gene sequences contain a C-terminally fused domain annotated as ctaG (not to be confused with the COX11/CtaG protein family), which is a transmembrane protein implicated in cytochrome c biogenesis in Bacillus species. 37,85,86 The presence of multiple copC genes in 37.6% of CopC-producing species may also support functional roles of CopC and CopD other than regulation (SI File 4). Copper imported by the CopCD system might also be used to metalate fully folded copper-containing proteins that are exported from the cytoplasm via the twin arginine translocation (TAT) pathway.…”

Extracytoplasmic Processes Impaired by Inactivation of trxA (Thioredoxin Gene) in Bacillus subtilis

“…SDS-PAGE was carried out using the Schägger and von Jagow buffer system (32). Isolation of membranes, N,N,NЈ,NЈ-tetramethyl-p-phenylenediamine (TMPD) oxidation activity staining of colonies on plates, and determination of cytochrome c oxidase activity were done as described previously (2). Staining of proteins for covalently bound heme after SDS-PAGE was carried out as described previously (36).…”