Aerobic expression of the cyf gene encoding cytochrome c-553 from Desulfovibrio vulgaris Hildenborough in Escherichia coli.
Monohaem cytochrome c-553 from Desulfovibrio vulgaris Hildenborough is encoded by the cyf gene which could be expressed in Escherichia coli to yield the periplasmic holoform of cytochrome c-553. Covalent haem attachment was shown by labelling with 5-amino[4-14C]laevulinic acid, the immediate precursor for haem biosynthesis. Visible-absorption spectroscopy demonstrated that the haem environment in the recombinant protein did not differ from the native protein. Optimal expression was obtained under aerobic conditions. Furthermore, efficient insertion of haem into a cytochrome c-553-beta-lactamase fusion protein could be demonstrated. We suggest that the varying success in heterologous expression of c-type cytochromes in E. coli may arise as a result of differences in the physical properties of the apoproteins.
Department of Biological Sciences, University of Calgary, Alberta, Canada.
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WB
Pollock
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Voordouw
D000332:Aerobiosis; D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D003574:Cytochrome c Group / Q000096:biosynthesis* / Q000235:genetics; D016969:Desulfovibrio vulgaris / Q000201:enzymology / Q000235:genetics*; D004790:Enzyme Induction; D004926:Escherichia coli; D005798:Genes, Bacterial; D006418:Heme / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics
D001426:Bacterial Proteins; D003574:Cytochrome c Group; D011993:Recombinant Fusion Proteins; C083951:cytochrome c553; D006418:Heme; C022302:cytochrome C-552