Abstract
The thiol-disulfide oxidoreductase DsbA carries out oxidative folding of extra-cytoplasmic proteins by catalyzing the formation of intramolecular disulfide bonds. It has an important role in various cellular functions, including cell division. The purple non-sulfur bacterium Rhodobacter capsulatus mutants lacking DsbA show severe temperature-sensitive and medium-dependent respiratory growth defects. In the presence of oxygen, at normal growth temperature (35°C), DsbA− mutants form colonies on minimal medium, but they do not grow on enriched medium where cells elongate and lyse. At lower temperatures (i.e., 25°C), cells lacking DsbA grow normally in both minimum and enriched media, however, they do not produce the cbb3-type cytochrome c oxidase (cbb3-Cox) on enriched medium. Availability of chemical oxidants (e.g., Cu2+ or a mixture of cysteine and cystine) in the medium becomes critical for growth and cbb3-Cox production in the absence of DsbA. Indeed, addition of Cu2+ to the enriched medium suppresses, and conversely, omission of Cu2+ from the minimal medium induces, growth and cbb3-Cox defects. Alleviation of these defects by addition of redox-active chemicals indicates that absence of DsbA perturbs cellular redox homeostasis required for the production of an active cbb3-Cox, especially in enriched medium where bioavailable Cu2+ is scarce. This is the first report describing that DsbA activity is required for full respiratory capability of R. capsulatus, and in particular, for proper biogenesis of its cbb3-Cox. We propose that absence of DsbA, besides impairing the maturation of the c-type cytochrome subunits, also affects the incorporation of Cu into the catalytic subunit of cbb3-Cox. Defective high affinity Cu acquisition pathway, which includes the MFS-type Cu importer CcoA, and lower production of the c-type cytochrome subunits lead together to improper assembly and degradation of cbb3-Cox.
Highlights
The Gram-negative, purple non-sulfur, facultative photosynthetic bacterium Rhodobacter capsulatus exhibits versatile growth modes, including anoxygenic photosynthesis (Ps) and aerobic respiration (Res) (Zannoni, 2004)
Cells growing by Res in enriched medium at 25◦C stopped dividing when shifted to 35◦C, yielding elongated cells that eventually lysed (Figure 2C, left side)
We have shown that R. capsulatus DsbA− mutants do not grow in enriched medium (MPYE) by Res they are proficient for Ps growth (Deshmukh et al, 2003)
Summary
The Gram-negative, purple non-sulfur, facultative photosynthetic bacterium Rhodobacter capsulatus exhibits versatile growth modes, including anoxygenic photosynthesis (Ps) and aerobic respiration (Res) (Zannoni, 2004). Mutants defective for cyt c maturation (Ccm) (i.e., covalent addition of heme to the apocyt c) (Sanders et al, 2010; Verissimo and Daldal, 2014) do not produce any c-type cyt (Sanders et al, 2010; Verissimo and Daldal, 2014), and are unable to grow by Ps, or by Res via the cbb3-Cox dependent branch They cannot catalyze the Nadi reaction (α-naphthol + dimethyl-phenylenediamine → indophenol blue + H2O) (Nadi− phenotype), which is a visual indicator for the cbb3-Cox activity (Koch et al, 1998). They can still grow by Res via the bd-Qox dependent branch, which contains no c-type cyt (Aygun-Sunar et al, 2006)
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