Engineering a prokaryotic apocytochrome c as an efficient substrate for Saccharomyces cerevisiae cytochrome c heme lyase.
Cytochromes c are heme proteins that require multiple maturation components, such as heme lyases, for cofactor incorporation. Saccharomyces cerevisiae has two heme lyases that are specific for apocytochromes c (CCHL) or c(1) (CC(1)HL). CCHL can covalently attach heme b groups to apocytochrome c substrates of eukaryotic but not prokaryotic origin. Besides their conserved Cys-Xxx-Xxx-Cys-His heme-binding motifs, the amino-terminal regions of apocytochrome c substrates appear to be important for CCHL function. In this study, we show for the first time that only two amino acid changes in the amino-terminal region of the non-CCHL substrate apocytochrome c(2) from Rhodobacter capsulatus are necessary and sufficient for efficient holocytochrome c formation by CCHL. This finding led us to propose a consensus sequence located at the amino-terminus of apocytochromes c, and critical for substrate recognition and heme ligation by CCHL.
Department of Biology, University of Pennsylvania, 433 South University Avenue, Philadelphia, PA 19104, USA.
Andreia F
AF
Verissimo
Joohee
J
Sanders
Fevzi
F
Daldal
Carsten
C
Sanders
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D016384:Consensus Sequence; D045304:Cytochromes c / Q000235:genetics / Q000378:metabolism*; D004252:DNA Mutational Analysis; D008190:Lyases / Q000378:metabolism*; D008969:Molecular Sequence Data; D015202:Protein Engineering; D016354:Rhodobacter capsulatus / Q000201:enzymology*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013379:Substrate Specificity
D001426:Bacterial Proteins; C014059:CYC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D045304:Cytochromes c; D008190:Lyases; C031581:cytochrome C synthetase