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9.B.14 The Putative Heme Handling Protein (HHP) Family


The proteins of the HHP family are large with ~650 amino acids and 15 or 16 putative TMSs. Parts of them are homologous to the E. coli CycZ putative heme exporter (9.B.14.2.1), the plant chloroplast cytochrome c biogenesis proteins such as CcsA of Chlamydomonas reinhardii (spP48269) and to HelC of Rhodobacter capsulatus (spP29961), also a 6 TMS protein thought to be involved in heme export. Ccl1 of R. capsulatus has been experimentally shown to have eleven TMSs. CcsA of Mycobacterium leprae has been shown to have 6 TMSs. UniProt puts these homologues in the CcsA/CcmF/CycK/Cel1 family. This family is distantly related to the UniProt CcmC/CycZ/HelC family (Lee et al., 2007). The functions of these proteins are not known, but some evidence suggests that at least some of them may be heme exporters (Baysse et al., 2003). CcmC of E. coli binds heme and interacts with CcmE, a heme chaperone protein that inserts heme into apocytochrome c (Ren and Thöny-Meyer, 2001). Kranz et al. (2009) have reviewed aspects of cytochrome c biogenesis including the mechanisms for covalent modifications and trafficking of heme, and for heme-iron redox control.

 Three major members of the HHP family (CcmC, CcmF and CcsBA), involved in cytochrome c biosynthesis, possess a conserved tryptophan-rich region (called the WWD domain) in an external loop at the inner membrane surface. The WWD domain binds heme to present it to an acceptor protein (apoCcmE for CcmC or apocytochrome c for CcmF and CcsBA) such that the heme vinyl group covalently attaches to the acceptor. CcmE only interacts stably with CcmC when heme is present. Endogenously synthesized heme enters the external WWD domain of CcmC either via a channel within this six-transmembrane-spanning protein or from the membrane (Richard-Fogal and Kranz, 2010).

Frawley and Kranz, (2009) showed that CcsBA exports and protects heme from oxidation. CcsBA has 10 apparent TMSs and reconstitutes cytochrome c synthesis in the E. coli periplasm; thus, CcsBA is a cytochrome c synthetase. Purified CcsBA contains heme in an 'external heme binding domain' for which two external histidines are shown to serve as axial ligands that protect the heme iron from oxidation. This may be the active site of the synthetase. Furthermore, two conserved histidines in TMSs are required for heme to travel to the external heme binding domain. Thus, CcsBA is a heme channel with a heme binding site within the bilayer. 

Organisms employ one of several different enzyme systems to mature cytochromes c (Simon and Hederstedt 2011). The biosynthetic process involves the periplasmic reduction of cysteine residues in the heme c attachment motif of the apocytochrome, transmembrane transport of heme b and stereospecific covalent heme attachment via thioether bonds. The biogenesis System II (or Ccs system) is employed by β-, δ- and ε-proteobacteria, Gram-positive bacteria, Aquificales and cyanobacteria, as well as by algal and plant chloroplasts. System II comprises four (sometimes only three) membrane-bound proteins: CcsA (or ResC) and CcsB (ResB) are the components of the cytochrome c synthase, whereas CcdA and CcsX (ResA) function in the generation of a reduced heme c attachment motif. Some ε-proteobacteria contain CcsBA fusion proteins constituting single polypeptide cytochrome c synthases especially amenable for functional studies (Simon and Hederstedt 2011).

References associated with 9.B.14 family:

Baert, B., C. Baysse, S. Matthijs, and P. Cornelis. (2008). Multiple phenotypic alterations caused by a c-type cytochrome maturation ccmC gene mutation in Pseudomonas aeruginosa. Microbiology 154: 127-138. 18174132
Baysse, C., S. Matthijs, M. Schobert, G. Layer, D. Jahn, and P. Cornelis. (2003). Co-ordination of iron acquisition, iron porphyrin chelation and iron-protoporphyrin export via the cytochrome c biogenesis protein CcmC in Pseudomonas fluorescens. Microbiology 149: 3543-3552. 14663086
Frawley, E.R. and R.G. Kranz. (2009). CcsBA is a cytochrome c synthetase that also functions in heme transport. Proc. Natl. Acad. Sci. USA 106: 10201-10206. 19509336
Goldman, B.S., D.L. Beck, E.M. Monika, and R.G. Kranz. (1998). Transmembrane heme delivery systems. Proc. Natl. Acad. Sci USA 95: 5003-5008. 9560218
Inoue, K., B.W. Dreyfuss, K.L. Kindle, D.B. Stern, S. Merchant, and O.A. Sodeinde. (1997). Ccs1, a nuclear gene required for the post-translational assembly of chloroplast c-type cytochromes. J. Biol. Chem. 272: 31747-31754. 9395519
Kranz, R.G., C. Richard-Fogal, J.S. Taylor, and E.R. Frawley. (2009). Cytochrome c biogenesis: mechanisms for covalent modifications and trafficking of heme and for heme-iron redox control. Microbiol. Mol. Biol. Rev. 73: 510-28, Table of Contents. 19721088
Lee, J.H., E.M. Harvat, J.M. Stevens, S.J. Ferguson, and M.H. Saier, Jr. (2007). Evolutionary origins of members of a superfamily of integral membrane cytochrome c biogenesis proteins. Biochim. Biophys. Acta. 1768: 2164-2181. 17706591
Page, M.D. and S.J. Ferguson. (1999). Mutational analysis of the Paracoccus denitrificans c-type cytochrome biosynthetic genes ccmABCDG: disruption of ccmC has distinct effects suggesting a role for CcmC independent of CcmAB. Microbiology 145: 3047-3057. 10589712
Page, M.D., Y. Sambongi, and S.J. Ferguson. (1998). Contrasting routes of c-type cytochrome assembly in mitochondria, chloroplasts and bacteria. Trends Biochem. Sci. 23: 103-108. 9581502
Pearce, D.A., M.D. Page, H.A.C. Norris, E.J. Tomlinson, and S.J. Ferguson. (1998). Identification of the contiguous Paracoccus denitrificans ccmF and ccmH genes: disruption of ccmF, encoding a putative transporter, results in formation of an unstable apocytochrome c and deficiency in siderophore production. Microbiology 144: 467-477. 9493384
Ren, Q. and L. Thony-Meyer. (2001). Physical interaction of CcmC with heme and the heme chaperone CcmE during cytochrome c maturation. J. Biol. Chem. 276: 32591-32596. 11384983
Richard-Fogal, C. and R.G. Kranz. (2010). The CcmC:heme:CcmE complex in heme trafficking and cytochrome c biosynthesis. J. Mol. Biol. 401: 350-362. 20599545
Schulz, H., E.C. Pellicioli, and L. Thöny-Meyer. (2000). New insights into the role of CcmC, CcmD and CcmE in the haem delivery pathway during cytochrome c maturation by a complete mutational analysis of the conserved tryptophan-rich motif of CcmC. Mol. Microbiol. 37: 1379-1388. 10998170
Schulz, H., R.A. Fabianek, E.C. Pellicioli, H. Hennecke, and L. Thöny-Meyer. (1999). Heme transfer to the heme chaperone CcmE during cytochrome c maturation requires the CcmC protein, which may function independently of the ABC-transporter CcmAB. Proc. Natl. Acad. Sci. USA 96: 6462-6467. 10339610
Simon, J. and L. Hederstedt. (2011). Composition and function of cytochrome c biogenesis System II. FEBS J. 278: 4179-4188. 21955752
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