Characterization of a cytochrome c gene located at the gene cluster for chlorate respiration in Ideonella dechloratans

Abstract

Anaerobic chlorate respiration requires electron transport from the bacterial inner membrane to the soluble periplasmic chlorate reductase. We have recently demonstrated that soluble c cytochromes function as electron carriers for chlorate reduction in Ideonella dechloratans (Smedja Bäcklund et al. 2009). In the present work, we describe a gene encoding soluble c-type cytochrome [cyt; GenBank ID: EU768872] located close to the gene cluster for chlorate reduction in I. dechloratans. The predicted amino acid sequence does not match any of the peptide masses or partial sequences obtained earlier from periplasmic c cytochromes. The gene, without the predicted signal sequence, was expressed heterologously in E. coli and the recombinant protein was purified, refolded and reconstituted with heme. The reconstituted protein shows spectral properties characteristic for c cytochromes, with an absorption maximum at 553 nm for the alpha band in the reduced state. Pyridine hemochrome analysis demonstrates the formation of covalently bound heme.

Structure and Evolution of Chlorate Reduction Composite Transposons

“…AJ296077.1) (29), a cytochrome c and molybdopterin-guanine dinucleotide biosynthesis gene (GenBank accession no. EU768872.1) (30), and clrABDC with the insertion sequence ISIde1 (GenBank accession no. AJ566363.1) (10) (Fig.…”

“…ACDC, NSS, PK, and AW-1 all contain a cytochrome c next to cld that is a possible candidate. However, in I. dechloratans , the cytochrome c in the chlorate reduction composite transposon does not donate electrons to ClrABC in vitro , suggesting that it does not provide the necessary link to the electron transport chain (30). Another cytochrome c in I. dechloratans has been shown to donate electrons to ClrABC in vitro (56), and we identified its sequence (see Fig.…”

Quinol-cytochrome c Oxidoreductase and Cytochrome c4 Mediate Electron Transfer during Selenate Respiration in Thauera selenatis

“…The authors suggest that the 6-kDa cytochrome might accept electrons from QCR, but no biochemical evidence for the involvement of QCR was reported. More recently, a cytochrome c gene located at the gene cluster for chlorate respiration in I. dechloratans has been cloned and overexpressed in E. coli but demonstrated not to function as an electron donor to purified chlorate reductase (49). It is interesting to speculate what the functions of the additional c-type cytochromes are in T. selenatis when grown on selenate.…”

Chlorate reductase is cotranscribed with cytochrome c and other downstream genes in the gene cluster for chlorate respiration of Ideonella dechloratans

“…The cld gene is located upstream of clrABDC, orientated in the opposite direction and separated from clrA by an insertion element (ISIde1). The cyc gene, which encodes a soluble c cytochrome, has the same direction as clrABDC and is found 126 bp downstream of clrC (Bohlin et al 2010). A similar arrangement of the cld and clrABDC genes was later established in Pseudomonas chloritidismutans AW-1 (GenBank GQ919187.1).…”

“…While the three key enzymes have been isolated and characterized in several strains (Nilsson, Rova and Smedja Bäcklund 2013), not much is known about the other components necessary for dissimilatory chlorate or perchlorate reduction, such as the participants of the electron transfer chains or regulatory factors. Ideonella dechloratans (Malmqvist et al 1994) was the first CRB to have its genes for chlorite dismutase (cld) and chlorate reductase (clrABDC) sequenced and these genes were found to be arranged in a gene cluster which later was shown to also include a cytochrome c gene, at first denoted cytc, here designated cyc (GenBank EU768872.1) and a molybdopterin-guanine dinucleotide biosynthesis gene (mobB) (Danielsson Thorell et al 2002;Danielsson Thorell et al 2003;Bohlin et al 2010) (Fig. 1).…”