Transmutation of a heme protein.
Residue Asn57 of bovine liver cytochrome b5 has been replaced with a cysteine residue, and the resulting variant has been isolated from recombinant Escherichia coli as a mixture of four major species: A, BI, BII, and C. A combination of electronic spectroscopy, 1H NMR spectroscopy, resonance Raman spectroscopy, electrospray mass spectrometry, and direct electrochemistry has been used to characterize these four major cytochrome derivatives. The red form A (E(m) = -19 mV) is found to possess a heme group bound covalently through a thioether linkage involving Cys57 and the alpha carbon of the heme 4-vinyl group. Form BI has a covalently bound heme group coupled through a thioether linkage involving the beta carbon of the heme 4-vinyl group. Form BII is similar to BI except that the sulfur involved in the thioether linkage is oxidized to a sulfoxide. The green form C (E(m) = 175 mV) possesses a noncovalently bound prosthetic group with spectroscopic properties characteristic of a chlorin. A mechanism is proposed for the generation of these derivatives, and the implications of these observations for the biosynthesis of cytochrome c and naturally occurring chlorin prosthetic groups are discussed.
Barker
P D
PD
Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, Canada.
Ferrer
J C
JC
Mylrajan
M
M
Loehr
T M
TM
Feng
R
R
Konishi
Y
Y
Funk
W D
WD
MacGillivray
R T
RT
Mauk
A G
AG
D000818:Animals; D002417:Cattle; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D015786:Cytochromes b5 / Q000737:chemistry* / Q000235:genetics*; D014644:Genetic Variation; D009682:Magnetic Resonance Spectroscopy; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D011524:Protoporphyrins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013053:Spectrophotometry; D013059:Spectrum Analysis, Raman
D010446:Peptide Fragments; D011524:Protoporphyrins; D011994:Recombinant Proteins; D015786:Cytochromes b5; C028025:protoporphyrin IX; D003545:Cysteine