Abstract
BackgroundIn the genome of H. pylori 26695, 149 proteins containing the CXXC motif characteristic of thioldisulfide oxidoreductases have been identified to date. However, only two of these proteins have a thioredoxin-like fold (i.e., HP0377 and HP0231) and are periplasm-located. We have previously shown that HP0231 is a dimeric oxidoreductase that catalyzes disulfide bond formation in the periplasm. Although HP0377 was originally described as DsbC homologue, its resolved structure and location of the hp0377 gene in the genome indicate that it is a counterpart of CcmG/DsbE.ResultsThe present work shows that HP0377 is present in H. pylori cells only in a reduced form and that absence of the main periplasmic oxidase HP0231 influences its redox state. Our biochemical analysis indicates that HP0377 is a specific reductase, as it does not reduce insulin. However, it possesses disulfide isomerase activity, as it catalyzes the refolding of scrambled RNase. Additionally, although its standard redox potential is -176 mV, it is the first described CcmG protein having an acidic pKa of the N-terminal cysteine of the CXXC motif, similar to E. coli DsbA or E. coli DsbC. The CcmG proteins that play a role in a cytochrome c-maturation, both in system I and system II, are kept in the reduced form by an integral membrane protein DsbD or its analogue, CcdA. In H. pylori HP0377 is re-reduced by CcdA (HP0265); however in E. coli it remains in the oxidized state as it does not interact with E. coli DsbD. Our in vivo work also suggests that both HP0377, which plays a role in apocytochrome reduction, and HP0378, which is involved in heme transport and its ligation into apocytochrome, provide essential functions in H. pylori.ConclusionsThe present data, in combination with the resolved three-dimensional structure of the HP0377, suggest that HP0377 is an unusual, multifunctional CcmG protein.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0471-z) contains supplementary material, which is available to authorized users.
Highlights
In the genome of H. pylori 26695, 149 proteins containing the CXXC motif characteristic of thioldisulfide oxidoreductases have been identified to date
The in vivo redox state of an oxidoreductase usually reflects its activity in the cell, i.e., proteins that function in the oxidizing pathway, such as EcDsbA, are maintained in the oxidized state in vivo, whereas proteins that function in the reducing pathway, such as CcmG, are maintained predominantly in the reduced state
We found that HP0377 is present in the reduced form in wild-type cells, which suggests that HP0377 functions as a reductase in H. pylori, which is in accord with its three dimensional structure [22]
Summary
In the genome of H. pylori 26695, 149 proteins containing the CXXC motif characteristic of thioldisulfide oxidoreductases have been identified to date. Only two of these proteins have a thioredoxin-like fold (i.e., HP0377 and HP0231) and are periplasm-located. We have previously shown that HP0231 is a dimeric oxidoreductase that catalyzes disulfide bond formation in the periplasm. In gram-negative bacteria, the oxidative protein folding takes place in the periplasm and is controlled by proteins from the Dsb family [1]. In the Roszczenko et al BMC Microbiology (2015) 15:135 and those contributing to reduction of a disulfide bond of the CXXCH heme-binding motifs [3,4,5]. The apocytochrome reduction is accomplished by the action of CcmG ( known as DsbE), CcmH, and DsbD or CcdA. DsbD/CcdA transfers electrons from cytoplasmic thioredoxin to CcmG. CcmG is reoxidized by shuttling its electrons to CcmH, which transfers them to apocytochrome c. The remaining proteins play a role in heme transport and its ligation to the apocytochrome [4]
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