Expression of doubly labeled Saccharomyces cerevisiae iso-1 ferricytochrome c and (1)H, (13)C and (15)N chemical shift assignments by multidimensional NMR.
We have expressed [U-(13)C,(15)N]-labeled Saccharomyces cerevisiae iso-1 cytochrome c C102T;K72A in Escherichia coli with a yield of 11 mg/l of growth medium. Nuclear magnetic resonance (NMR) studies were conducted on the Fe(3+) form of the protein. We report herein chemical shift assignments for amide (1)H and (15)N, (13)C(omicron), (13)C(alpha), (13)C(beta), (1)H(alpha) and (1)H(beta) resonances based upon a series of three-dimensional NMR experiments: HNCA, HN(CO)CA, HNCO, HN(CA)CO, HNCACB, HCA(CO)N, HCCH-TOCSY and HBHA(CBCA)NH. An investigation of the chemical shifts of the threonine residues was also made by using density functional theory in order to help solve discrepancies between (15)N chemical shift assignments reported in this study and those reported previously.
Szabo
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Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, IL 61801, USA.
Sanders
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Chien
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Oldfield
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D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D003574:Cytochrome c Group / Q000737:chemistry*; D045304:Cytochromes c; D006859:Hydrogen; D007527:Isoenzymes / Q000737:chemistry; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013912:Threonine / Q000737:chemistry
C014059:CYC1 protein, S cerevisiae; D002247:Carbon Isotopes; D003574:Cytochrome c Group; D007527:Isoenzymes; D009587:Nitrogen Isotopes; D029701:Saccharomyces cerevisiae Proteins; D013912:Threonine; D006859:Hydrogen; D045304:Cytochromes c