Cloning and sequence of cymA, a gene encoding a tetraheme cytochrome c required for reduction of iron(III), fumarate, and nitrate by Shewanella putrefaciens MR-1

Abstract

The cymA gene, which encodes a tetraheme cytochrome c, was cloned from Shewanella putrefaciens MR-1. This gene complemented a mutant which had a TnphoA insertion in cymA and which was deficient in the respiratory reduction of iron(III), nitrate, fumarate, and manganese(IV). The 561-bp nucleotide sequence of cymA encodes a protein of 187 amino acids with a predicted molecular mass of 20.8 kDa. No N-terminal signal sequence was readily apparent; consistent with this, a cytochrome with a size of 21 kDa was detected in the wild type but was absent in the insertional mutant. The cymA gene is transcribed into an mRNA; the major transcript was approximately 790 bases, suggesting that it is not part of a multicistronic operon. This RNA transcript was not detected in the cymA mutant. The CymA protein was found in the cytoplasmic membrane and soluble fraction of MR-1, and it shares partial amino acid sequence homology with multiheme c-type cytochromes from other bacteria. These cytochromes are ostensibly involved in the transfer of electrons from the cytoplasmic membrane to acceptors in the periplasm. The localization of the fumarate and iron(III) reductases to the periplasm and outer membrane of MR-1, respectively, suggests the possibility of a similar electron transfer role for CymA.

Reductive activation of hexavalent chromium by human lung epithelial cells: Generation of Cr(V) and Cr(V)-thiol species

“…For protein analysis, the TCA-insoluble pellets (from above) were dissolved in 0.1 M NaOH for ≥5 days at room temperature and then protein was determined by a modified Lowry method, with bovine serum albumin as the standard [43].…”

A dynamic periplasmic electron transfer network enables respiratory flexibility beyond a thermodynamic regulatory regime

“…Moreover, the majority of pathways are dependent on the catalysis of electron transfer into the periplasm by the menaquinol oxidase and tetraheme cytochrome CymA . Unsurprisingly, a cymA-deficient mutant shows growth deficiencies with a variety of anoxic electron acceptors (Myers and Myers, 1997). Although it was shown experimentally in vivo and in vitro that CymA can directly transfer electrons to the respiratory fumarate reductase FccA, it was suggested that a direct transfer would also be in place toward the periplasmic nitrate and nitrite reductases NapAB and NrfA, respectively (Schwalb et al, 2002;Gao et al, 2009;Schuetz et al, 2009).…”

The periplasmic nitrate reductase in Shewanella: the resolution, distribution and functional implications of two NAP isoforms, NapEDABC and NapDAGHB

“…The sequence identity values for the other subunits of the NAP-a and NAP-b isoforms are given in Table 3. Based upon experimental work with S. denitrificans (NAP-a-based system) (Brettar et al, 2002) and S. oneidensis MR-1 (NAP-b-based-system) (Cruz-Garcia et al, 2007;Myers & Myers, 1997;Schwalb et al, 2002), both NAP isoforms must be catalytically active. It may be that in species of Shewanella that contain both NAP-a and NAP-b, the NAP isoforms are differentially regulated under different environmental conditions (pH, oxygen availability, quinol pools).…”

“…The nap operons of the e-proteobacteria Wolinella succinogenes (napAGHBFLD) (Kern & Simon, 2008;Simon et al, 2003) and Campylobacter jejuni (napAGHBLD) do not include the napC gene. The function of NapC in NAP in these species and in S. oneidensis MR-1, which also lacks NapC, may be met by the periplasmic c-type cytochrome homologue CymA, which is also from the NapC/NirT family (Gao et al, 2009;Murphy & Saltikov, 2007;Myers & Myers, 1997, 2000Schwalb et al, 2003). The napCMADGH operon of the d-proteobacterium Desulfovibrio desulfuricans lacks the napB gene (Marietou et al, 2005).…”