title: Probing why trypanosomes assemble atypical cytochrome c with an AxxCH haem-binding motif instead of CxxCH.
abstract: Mitochondrial cytochromes c and c1 are core components of the respiratory chain of all oxygen-respiring eukaryotes. These proteins contain haem, covalently bound to the polypeptide in a catalysed post-translational modification. In all eukaryotes, except members of the protist phylum Euglenozoa, haem attachment is to the cysteine residues of a CxxCH haem-binding motif. In the Euglenozoa, which include medically relevant trypanosomatid parasites, haem attachment is to a single cysteine residue in an AxxCH haem-binding motif. Moreover, genes encoding known c-type cytochrome biogenesis machineries are all absent from trypanosomatid genomes, indicating the presence of a novel biosynthetic apparatus. In the present study, we investigate expression and maturation of cytochrome c with a typical CxxCH haem-binding motif in the trypanosomatids Crithidia fasciculata and Trypanosoma brucei. Haem became attached to both cysteine residues of the haem-binding motif, indicating that, in contrast with previous hypotheses, nothing prevents formation of a CxxCH cytochrome c in euglenozoan mitochondria. The cytochrome variant was also able to replace the function of wild-type cytochrome c in T. brucei. However, the haem attachment to protein was not via the stereospecifically conserved linkage universally observed in natural c-type cytochromes, suggesting that the trypanosome cytochrome c biogenesis machinery recognized and processed only the wild-type single-cysteine haem-binding motif. Moreover, the presence of the CxxCH cytochrome c resulted in a fitness cost in respiration. The level of cytochrome c biogenesis in trypanosomatids was also found to be limited, with the cells operating at close to maximum capacity.
authors.affiliation: Faculty of Health and Medicine, Division of Biomedical and Life Sciences, Lancaster University, Lancaster LA1 4YQ, UK.
authors.forename: Michael L
authors.initials: ML
authors.lastname: Ginger
authors.forename: Katharine A
authors.initials: KA
authors.lastname: Sam
authors.forename: James W A
authors.initials: JW
authors.lastname: Allen
mesh_terms: D020816:Amino Acid Motifs; D001483:Base Sequence; D001665:Binding Sites; D016831:Crithidia fasciculata / Q000235:genetics / Q000378:metabolism*; D045304:Cytochromes c / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004579:Electron Transport; D019143:Evolution, Molecular; D006418:Heme / Q000737:chemistry; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D006418:Heme; D045304:Cytochromes c