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Structural and functional characterization of CcmG from Pseudomonas aeruginosa, a key component of the bacterial cytochrome c maturation apparatus
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
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2010 (English)In: Proteins: Structure, Function, and Genetics, ISSN 0887-3585, E-ISSN 1097-0134, ISSN 20544959, Vol. 78, no 10, 2213-2221 p.Article in journal (Refereed) Published
Abstract [en]

The cytochrome c maturation process is carried out in the bacterial periplasm, where some specialized thiol-disulfide oxidoreductases work in close synergy for the correct reduction of oxidized apocytochrome before covalent heme attachment. We present a structural and functional characterization of the soluble periplasmic domain of CcmG from the opportunistic pathogen P. aeruginosa (Pa-CcmG), a component of the protein machinery involved in cyt c maturation in gram-negative bacteria. X-ray crystallography reveals that Pa-CcmG is a TRX-like protein; high-resolution crystal structures show that the oxidized and the reduced forms of the enzyme are identical except for the active-site disulfide. The standard redox potential was calculated to be E-0' = -0.213 V at pH 7.0; the pK(a) of the active site thiols were pK(a) = 6.13 +/- 0.05 for the N-terminal Cys74 and pK(a) = 10.5 +/- 0.17 for the C-terminal Cys77. Experiments were carried out to characterize and isolate the mixed disulfide complex between Pa-CcmG and Pa-CcmH (the other redox active component of System I in P. aeruginosa). Our data indicate that the target disulfide of this TRX-like protein is not the intramolecular disulfide of oxidized Pa-CcmH, but the intermolecular disulfide formed between Cys28 of Pa-CcmH and DTNB used for the in vitro experiments. This observation suggests that, in vivo, the physiological substrate of Pa-CcmG may be the mixed-disulfide complex between Pa-CcmH and apo-cyt. Proteins 2010; 78:2213-2221. (C) 2010 Wiley-Liss, Inc.

Place, publisher, year, edition, pages
2010. Vol. 78, no 10, 2213-2221 p.
Keyword [en]
thiol-disulfide oxidoreductases, thioredoxin-like protein, pKa, DTNB, cytocrome c biogenesis, cytochrome c heme binding motif
National Category
Medical and Health Sciences Natural Sciences
URN: urn:nbn:se:uu:diva-136010DOI: 10.1002/prot.22733ISI: 000279387400003OAI: oai:DiVA.org:uu-136010DiVA: diva2:376242
Available from: 2010-12-10 Created: 2010-12-09 Last updated: 2011-06-28Bibliographically approved

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Jemth, Per
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