Reduction of the periplasmic disulfide bond isomerase, DsbC, occurs by passage of electrons from cytoplasmic thioredoxin

Abstract

The Escherichia coli periplasmic protein DsbC is active both in vivo and in vitro as a protein disulfide isomerase. For DsbC to attack incorrectly formed disulfide bonds in substrate proteins, its two active-site cysteines should be in the reduced form. Here we present evidence that, in wild-type cells, these two cysteines are reduced. Further, we show that a pathway involving the cytoplasmic proteins thioredoxin reductase and thioredoxin and the cytoplasmic membrane protein DsbD is responsible for the reduction of these cysteines. Thus, reducing potential is passed from cytoplasmic electron donors through the cytoplasmic membrane to DsbC. This pathway does not appear to utilize the cytoplasmic glutathione-glutaredoxin pathway. The redox state of the active-site cysteines of DsbC correlates quite closely with its ability to assist in the folding of proteins with multiple disulfide bonds. Analysis of the activity of mutant forms of DsbC in which either or both of these cysteines have been altered further supports the role of DsbC as a disulfide bond isomerase.

Genomic island genes in a coastal marine Synechococcus strain confer enhanced tolerance to copper and oxidative stress

“…In E. coli, DsbD is a inner membrane-bound protein that transfers electrons to DsbC, a disulfide isomerase that repairs nonnative disulfide bonds in the periplasm (Rietsch et al, 1997) and DsbE, a thioredoxin protein essential to cytochrome c maturation (Stirnimann et al, 2005). The DsbD knockout is extremely copper sensitive, which is not surprising, since copper as a redox metal catalyzes disulfide bonds quite readily (Katzen and Beckwith, 2003), and the entire pathway is thought to be involved in copper resistance and potentially other oxidative stressors (Hiniker et al, 2005).…”

Thioredoxin 2, an Oxidative Stress-induced Protein, Contains a High Affinity Zinc Binding Site

“…The first one, TrxA, has been identified 40 years ago and is well characterized. It is a 12 kDa protein, which serves as electron donor for 3Ј-phosphoadenosyl sulfate (PAPS) 1 reductase, ribonucleotide reductase, methionine sulfoxide reductase, and the membrane protein DsbD (2,4). It is also an essential subunit of the bacteriophage T7 DNA polymerase.…”

SoxV transfers electrons to the periplasm of Paracoccus pantotrophus – an essential reaction for chemotrophic sulfur oxidation

“…Since the direction of the electron transfer from the cytoplasm to the periplasm appeared to be identical with that of the DsbD system of E. coli (Rietsch et al, 1997;Stewart et al, 1999;Chung et al, 2000;Katzen & Beckwith, 2000) the effect of reductants on the thiosulfateoxidizing activity of whole cells of the mutant strain was examined. Indeed, addition of reductant (1 mM DTT) to mixotrophically cultivated strain GBVV in the early stationary phase resulted in a 50-fold increase in formation of the thiosulfate-oxidizing activity of whole cells (Fig.…”

“…Members of the DsbD and CcdA families are involved in the transport of electrons from the cytoplasm to the periplasm for reduction of apocytochromes to enable addition of the haem moiety (reviewed by Thöny-Meyer, 1997). The electrons are transferred from a cytoplasmic thioredoxin first to the central transmembrane b-domain of DsbD then to the Cterminal thioredoxin-like c-domain and therefrom to the N-terminal a-domain, which then reduces either DsbC or CcmG (Rietsch et al, 1997;Stewart et al, 1999;Chung et al, 2000;Katzen & Beckwith, 2000).…”

“…Members of the DsbD and CcdA families are involved in the transport of electrons from the cytoplasm to the periplasm for reduction of apocytochromes to enable addition of the haem moiety (reviewed by Thöny-Meyer, 1997). The electrons are transferred from a cytoplasmic thioredoxin first to the central transmembrane b-domain of DsbD then to the Cterminal thioredoxin-like c-domain and therefrom to the N-terminal a-domain, which then reduces either DsbC or CcmG (Rietsch et al, 1997;Stewart et al, 1999;Chung et al, 2000;Katzen & Beckwith, 2000).The Gram-negative, facultatively chemolithoautotrophic bacterium Paracoccus pantotrophus grows under aerobic conditions lithotrophically with molecular hydrogen and thiosulfate as energy source for autotrophic carbon dioxide fixation (Robertson & Kuenen, 1983;Rainey et al, 1999). In P. pantotrophus the soxVWXYZABCDEFGH genes constitute two transcriptional units, soxVW and soxX-H (Rother et al, 2005).…”