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. 1996 Nov;71(5):2848–2856. doi: 10.1016/S0006-3495(96)79481-X

Assignment of 1H and 13C hyperfine-shifted resonances for tuna ferricytochrome c.

S F Sukits 1, J D Satterlee 1
PMCID: PMC1233771  PMID: 8913622

Abstract

Tuna ferricytochrome c has been used to demonstrate the potential for completely assigning 1H and 13C strongly hyperfine-shifted resonances in metalloprotein paramagnetic centers. This was done by implementation of standard two-dimensional NMR experiments adapted to take advantage of the enhanced relaxation rates of strongly hyperfine-shifted nuclei. The results show that complete proton assignments of the heme and axial ligands can be achieved, and that assignments of several strongly shifted protons from amino acids located close to the heme can also be made. Virtually all proton-bearing heme 13C resonances have been located, and additional 13C resonances from heme vicinity amino acids are also identified. These results represent an improvement over previous proton resonance assignment efforts that were predicated on the knowledge of specific assignments in the diamagnetic protein and relied on magnetization transfer experiments in heterogeneous solutions composed of mixtures of diamagnetic ferrocytochrome c and paramagnetic ferricytochrome c. Even with that more complicated procedure, complete heme proton assignments for ferricytochrome c have never been demonstrated by a single laboratory. The results presented here were achieved using a more generally applicable strategy with a solution of the uniformly oxidized protein, thereby eliminating the requirement of fast electron self-exchange, which is a condition that is frequently not met.

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Selected References

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  1. Alam S. L., Satterlee J. D. Complete heme proton hyperfine resonance assignments of the Glycera dibranchiata component IV metcyano monomer hemoglobin. Biochemistry. 1994 Apr 5;33(13):4008–4018. doi: 10.1021/bi00179a030. [DOI] [PubMed] [Google Scholar]
  2. Bernstein F. C., Koetzle T. F., Williams G. J., Meyer E. F., Jr, Brice M. D., Rodgers J. R., Kennard O., Shimanouchi T., Tasumi M. The Protein Data Bank: a computer-based archival file for macromolecular structures. J Mol Biol. 1977 May 25;112(3):535–542. doi: 10.1016/s0022-2836(77)80200-3. [DOI] [PubMed] [Google Scholar]
  3. Feng Y. Q., Roder H., Englander S. W. Assignment of paramagnetically shifted resonances in the 1H NMR spectrum of horse ferricytochrome c. Biophys J. 1990 Jan;57(1):15–22. doi: 10.1016/S0006-3495(90)82502-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Feng Y., Roder H., Englander S. W., Wand A. J., Di Stefano D. L. Proton resonance assignments of horse ferricytochrome c. Biochemistry. 1989 Jan 10;28(1):195–203. doi: 10.1021/bi00427a027. [DOI] [PubMed] [Google Scholar]
  5. Gao Y., Boyd J., Pielak G. J., Williams R. J. Proton nuclear magnetic resonance as a probe of differences in structure between the C102T and F82S,C102T variants of iso-1-cytochrome c from the yeast Saccharomyces cerevisiae. Biochemistry. 1991 Jul 16;30(28):7033–7040. doi: 10.1021/bi00242a032. [DOI] [PubMed] [Google Scholar]
  6. Gao Y., Boyd J., Williams R. J., Pielak G. J. Assignment of proton resonances, identification of secondary structural elements, and analysis of backbone chemical shifts for the C102T variant of yeast iso-1-cytochrome c and horse cytochrome c. Biochemistry. 1990 Jul 31;29(30):6994–7003. doi: 10.1021/bi00482a007. [DOI] [PubMed] [Google Scholar]
  7. Keller R. M., Wüthrich K. Assignment of the heme c resonances in the 360 MHz H NMR spectra of cytochrome c. Biochim Biophys Acta. 1978 Mar 28;533(1):195–208. doi: 10.1016/0005-2795(78)90564-0. [DOI] [PubMed] [Google Scholar]
  8. Moench S. J., Satterlee J. D. Proton NMR comparison of the Saccharomyces cerevisiae ferricytochrome c isozyme-1 monomer and covalent disulfide dimer. J Biol Chem. 1989 Jun 15;264(17):9923–9931. [PubMed] [Google Scholar]
  9. Moench S. J., Shi T. M., Satterlee J. D. Proton-NMR studies of the effects of ionic strength and pH on the hyperfine-shifted resonances and phenylalanine-82 environment of three species of mitochondrial ferricytochrome c. Eur J Biochem. 1991 May 8;197(3):631–641. doi: 10.1111/j.1432-1033.1991.tb15953.x. [DOI] [PubMed] [Google Scholar]
  10. Moore G. R., Williams R. J. The solution structures of tuna and horse cytochromes c. Eur J Biochem. 1980 Feb;103(3):533–541. doi: 10.1111/j.1432-1033.1980.tb05977.x. [DOI] [PubMed] [Google Scholar]
  11. Moss G. P. Nomenclature of tetrapyrroles. Recommendations 1986 IUPAC-IUB Joint Commission on Biochemical Nomenclature (JCBN). Eur J Biochem. 1988 Dec 15;178(2):277–328. doi: 10.1111/j.1432-1033.1988.tb14453.x. [DOI] [PubMed] [Google Scholar]
  12. Oh B. H., Mooberry E. S., Markley J. L. Multinuclear magnetic resonance studies of the 2Fe.2S* ferredoxin from Anabaena species strain PCC 7120. 2. Sequence-specific carbon-13 and nitrogen-15 resonance assignments of the oxidized form. Biochemistry. 1990 Apr 24;29(16):4004–4011. doi: 10.1021/bi00468a030. [DOI] [PubMed] [Google Scholar]
  13. Qin J., La Mar G. N. Complete sequence-specific 1H NMR resonance assignment of hyperfine-shifted residues in the active site of a paramagnetic protein: application to Aplysia cyano-metmyoglobin. J Biomol NMR. 1992 Nov;2(6):597–618. doi: 10.1007/BF02192849. [DOI] [PubMed] [Google Scholar]
  14. Santos H., Turner D. L. 13C and proton NMR studies of horse cytochrome c. Assignment and temperature dependence of methyl resonances. FEBS Lett. 1986 Jan 1;194(1):73–77. doi: 10.1016/0014-5793(86)80054-0. [DOI] [PubMed] [Google Scholar]
  15. Santos H., Turner D. L. 13C and proton NMR studies of horse cytochrome c. Systematic assignment of methyl and methine resonances in both oxidation states. Eur J Biochem. 1992 Jun 15;206(3):721–728. doi: 10.1111/j.1432-1033.1992.tb16978.x. [DOI] [PubMed] [Google Scholar]
  16. Santos H., Turner D. L. Proton NMR studies of horse ferricytochrome c. Completion of the assignment of the well resolved hyperfine shifted resonances. FEBS Lett. 1987 Dec 21;226(1):179–185. doi: 10.1016/0014-5793(87)80575-6. [DOI] [PubMed] [Google Scholar]
  17. Satterlee J. D., Moench S. Proton hyperfine resonance assignments using the nuclear Overhauser effect for ferric forms of horse and tuna cytochrome c. Biophys J. 1987 Jul;52(1):101–107. doi: 10.1016/S0006-3495(87)83193-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Takano T., Dickerson R. E. Conformation change of cytochrome c. II. Ferricytochrome c refinement at 1.8 A and comparison with the ferrocytochrome structure. J Mol Biol. 1981 Nov 25;153(1):95–115. doi: 10.1016/0022-2836(81)90529-5. [DOI] [PubMed] [Google Scholar]
  19. Williams G., Moore G. R., Porteous R., Robinson M. N., Soffe N., Williams R. J. Solution structure of mitochondrial cytochrome c. I. 1H nuclear magnetic resonance of ferricytochrome c. J Mol Biol. 1985 Jun 5;183(3):409–428. doi: 10.1016/0022-2836(85)90011-7. [DOI] [PubMed] [Google Scholar]
  20. Yamamoto Y., Nanai N., Inoue Y., Chûjô R. Natural abundance 13C-NMR study of paramagnetic horse heart ferricytochrome c cyanide complex: assignment of hyperfine shifted heme methyl carbon resonances. Biochem Biophys Res Commun. 1988 Feb 29;151(1):262–269. doi: 10.1016/0006-291x(88)90588-8. [DOI] [PubMed] [Google Scholar]
  21. de Ropp J. S., Yu L. P., La Mar G. N. 2D NMR of paramagnetic metalloenzymes: cyanide-inhibited horseradish peroxidase. J Biomol NMR. 1991 Jul;1(2):175–190. doi: 10.1007/BF01877229. [DOI] [PubMed] [Google Scholar]

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