Oxygen-17 Nuclear Magnetic Resonance Spectroscopic Studies of Carbonmonoxy Hemoproteins

Hee Cheon Lee, Eric Oldfield

Research output: Contribution to journalArticlepeer-review

Abstract

We have obtained surprisingly narrow 17O nuclear magnetic resonance (NMR) spectra at 8.45 and 11.7 T (corresponding to 17O frequencies of 48.8 and 67.8 MHz) from C17O ligands bound to aqueous ferrous myoglobin from Physeter catodon (sperm whale MbCO), from adult human ferrous hemoglobin (HbCO A), and from ferrous hemoglobin from Oryctolagus cuniculus (rabbit HbCO). The 17O NMR signals from these hemoproteins are not only narrower than anticipated, but in the case of sperm whale MbCO the line shape is distinctly non-Lorentzian. We have thus used the dispersion versus absorption (DISPA) plot method to investigate the origin of these unusual line widths and line shapes and demonstrate that they originate from the multiexponential nature of quadrupolar relaxation outside of the “extreme-narrowing” limit (ω0τC> 1). We find from the DISPA analysis and from spin-lattice relaxation time (T1) measurements that the 17O nuclear quadrupole coupling constant (QCC) for sperm whale MbCO is 0.95 MHz, and the rotational correlation time, τc, is 14 ns (at ω0τC= 5.8). This indicates a rigid heme-CO unit in sperm whale MbCO, Applying the same type of analysis to human HbCO yields 17O QCC values of 0.9 MHz and τc of 23 ns (at ω0τc = 10). In all cases, our results are consistent with an 17O chemical shift anisotropy (σ- σ) value of about 800 ppm for the CO ligand. These results are important for several reasons: first, they represent the first observation of high-resolution 170 NMR spectra of the CO ligands in metalloproteins. Second, they represent the first experimental demonstration of multiexponential relaxation of a spin I = 5/2 nucleus and its complete analysis with relaxation theory. Third, our results on sperm whale MbCO, taken together with 13C NMR relaxation data, indicate little “internal motion” of the heme-CO group in this system. Our results also demonstrate a linear relationship between the 17O NMR chemical shift and vco, the infrared stretching frequency of the CO ligand, and between the 17O chemical shift and the CO binding affinity of the protein. In addition, the 17O NMR results are also in good agreement with previous time-differential perturbed γ-ray angular correlation (PAC) results on [111In]myglobin and -hemoglobin (Marshall, A. G.; Lee, K. M.; Martin, P. W. J. Am. Chem. Soc. 1980, 102, 1460), and some molecular interpretations of the NMR and PAC results are offered.

Original languageEnglish (US)
Pages (from-to)1584-1590
Number of pages7
JournalJournal of the American Chemical Society
Volume111
Issue number5
DOIs
StatePublished - Mar 1989

ASJC Scopus subject areas

  • Catalysis
  • General Chemistry
  • Biochemistry
  • Colloid and Surface Chemistry

Fingerprint

Dive into the research topics of 'Oxygen-17 Nuclear Magnetic Resonance Spectroscopic Studies of Carbonmonoxy Hemoproteins'. Together they form a unique fingerprint.

Cite this