1887

Abstract

Bacterial cytochrome maturation occurs at the outside of the cytoplasmic membrane, requires transport of haem across the membrane, and depends on membrane-bound cytochrome haem lyase (CCHL), an enzyme that catalyses covalent attachment of haem to apocytochrome . Epsilonproteobacteria such as use the cytochrome biogenesis system II and contain unusually large CCHL proteins of about 900 amino acid residues that appear to be fusions of the CcsB and CcsA proteins found in other bacteria. CcsBA-type CCHLs have been proposed to act as haem transporters that contain two haem coordination sites located at different sides of the membrane and formed by histidine pairs. cells contain three CcsBA-type CCHL isoenzymes (NrfI, CcsA1 and CcsA2) that are known to differ in their specificity for apocytochromes and apparently recognize different haem binding motifs such as CXCH (by CcsA2), CXCK (by NrfI) and CXCH (by CcsA1). In this study, conserved histidine residues were individually replaced by alanine in each of the CCHLs. Characterization of NrfI and CcsA1 variants in demonstrated that a set of four histidines is essential for maturing the dedicated multihaem cytochromes NrfA and MccA, respectively. The function of CcsA2 variants produced in was also found to depend on each of these four conserved histidine residues. The presence of imidazole in the growth medium of both and rescued the cytochrome biogenesis activity of most histidine variants, albeit to different extents, thereby implying the presence of two functionally distinct histidine pairs in each CCHL. The data support a model in which two conserved haem binding sites are involved in haem transport catalysed by CcsBA-type CCHLs.

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2010-12-01
2019-12-07
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Nucleotide primers used to modify CCHL-encoding genes [PDF](45 KB)

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Full-length alignment of selected epsilonproteobacterial CCHL primary structures [PDF](43 KB)

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