Sequential main-chain proton and carbon resonance assignments for wild-type yeast iso-1 ferrocytochrome c and identification of secondary structural elements.
Yeast iso-1 cytochrome c is a naturally occurring protein that possesses an unusually reactive Cys102 that imbues iso-1 with a complicated solution chemistry which includes spontaneous dimerization and poorly characterized redox reactions. For this reason previous studies of this typical member of the c-type cytochromes have been relegated to variant proteins in which the 102 position has been mutated, with most common changes involving serine and threonine. However, we have determined sequential 1H resonance assignments for the wild-type native protein because it is the actual participant in yeast mitochondrial electron transfer processes and because the wild-type native protein should be the fundamental assignment basis. In addition to 1H resonance assignments for 97 of 106 amino acids, we have also provided an extensive database of long-range NOEs. Comparison of these NOEs and a chemical shift index based upon alpha-H resonances has lead to identification of solution secondary structural elements that are consistent with the solid-state crystal structure. Although there is currently no efficient expression system that would facilitate isotope labeling of iso-1 cytochrome c, we tried to assess the usefulness of future heteronuclear experiments by using natural-abundance 1H/13C HMQC experiments to unambiguously assign 35 alpha-C resonances.
Sukits
S F
SF
Department of Chemistry, Washington State University, Pullman 99164-4630, USA.
Satterlee
J D
JD
D000595:Amino Acid Sequence; D002244:Carbon / Q000032:analysis; D003574:Cytochrome c Group / Q000737:chemistry*; D045304:Cytochromes c; D006859:Hydrogen / Q000032:analysis; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins; D017421:Sequence Analysis
C014059:CYC1 protein, S cerevisiae; D003574:Cytochrome c Group; D029701:Saccharomyces cerevisiae Proteins; D002244:Carbon; D006859:Hydrogen; D045304:Cytochromes c