Goddard, Alan D. and Stevens, Julie M. and Rondelet, Arnaud and Nomerotskaia, Elena and Allen, James W. A. and Ferguson, Stuart J. (2010) Comparing the substrate specificities of cytochrome c biogenesis Systems I and II. FEBS Journal, 277 (3). pp. 726-737. ISSN 1742-464X

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Abstract

c-Type cytochromes require specific post-translational protein systems, which vary in different organisms, for the characteristic covalent attachment of heme to the cytochrome polypeptide. Cytochrome c biogenesis System II, found in chloroplasts and many bacteria, comprises four subunits, two of which (ResB and ResC) are the minimal functional unit. The ycf5 gene from Helicobacter pylori encodes a fusion of ResB and ResC. Heterologous expression of ResBC in Escherichia coli lacking its own biogenesis machinery allowed us to investigate the substrate specificity of System II. ResBC is able to attach heme to monoheme c-type cytochromes c 550 from Paracoccus denitrificans and c 552 from Hydrogenobacter thermophilus, both normally matured by System I. The production of holocytochrome is enhanced by the addition of exogenous reductant. Single-cysteine variants of these cytochromes were not efficiently matured by System II, but System I was able to produce detectable amounts of AXXCH variants; this adds to evidence that there is no obligate requirement for a disulfide-bonded intermediate for the latter c-type cytochrome biogenesis system. In addition, System II was able to mature an AXXAH-containing variant into a b-type cytochrome, with implications for both heme supply to the periplasm and substrate recognition by System II.

Item Type:	Article
Additional Information:	c-Type cytochromes require specific post-translational protein systems, which vary in different organisms, for the characteristic covalent attachment of heme to the cytochrome polypeptide. Cytochrome c biogenesis System II, found in chloroplasts and many bacteria, comprises four subunits, two of which (ResB and ResC) are the minimal functional unit. The ycf5 gene from Helicobacter pylori encodes a fusion of ResB and ResC. Heterologous expression of ResBC in Escherichia coli lacking its own biogenesis machinery allowed us to investigate the substrate specificity of System II. ResBC is able to attach heme to monoheme c-type cytochromes c 550 from Paracoccus denitrificans and c 552 from Hydrogenobacter thermophilus, both normally matured by System I. The production of holocytochrome is enhanced by the addition of exogenous reductant. Single-cysteine variants of these cytochromes were not efficiently matured by System II, but System I was able to produce detectable amounts of AXXCH variants; this adds to evidence that there is no obligate requirement for a disulfide-bonded intermediate for the latter c-type cytochrome biogenesis system. In addition, System II was able to mature an AXXAH-containing variant into a b-type cytochrome, with implications for both heme supply to the periplasm and substrate recognition by System II.
Keywords:	Cytochrome c, Cytochrome c maturation, Heme, Heme provision, System II
Subjects:	C Biological Sciences > C700 Molecular Biology, Biophysics and Biochemistry
Divisions:	College of Sciences > Faculty of Science > School of Life Sciences
Depositing User:	Alan Goddard
Date Deposited:	16 Nov 2012 08:58
Last Modified:	16 Nov 2012 08:58
URI:	http://eprints.lincoln.ac.uk/id/eprint/6837

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