Victor Glushko

Expressions are presented for the nuclear Overhauser enhancement (NOE), the spin-lattice relaxation time (T1), and the spin-spin relaxation time (T2) of a 13C nucleus relaxing by a dipolar interaction with one proton under conditions of complete proton decoupling, and without assuming that the extreme narrowing limit applies. Specific equations are derived for a C–H group in a rigid molecule rotating isotropically and also for a C–H group with one degree of internal motion attached to a molecule undergoing isotropic rotational reorientation. Numerical results are presented for T1, T2, and the NOE of a C–H group in a rigid molecule (undergoing purely dipolar relaxation) as a function of the rotational correlation time (τR) and the resonance frequency (ωC). T1 goes through a minimum when τRωC ≈ 0.8. The NOE varies from the expected value of 2.988 in the extreme narrowing limit to 1.153 when 1/τR is much smaller than the resonance frequency. The numerical results indicate that the signal-to-noise ratio in proton-decoupled 13C Fourier transform spectra of macromolecules in solution may not improve significantly by going to very high magnetic field strengths (such as 51.7 kG), because the increase in basic sensitivity can be offset by a decrease in the NOE and the T2/T1 ratio. The magnitude of the latter two parameters is strongly dependent on τR and the magnetic field strength. Numerical results are also presented for a C–H group with one degree of internal motion. 1/T2 is a monotonically increasing function of τR and τG (the correlation time for internal rotation). The NOE and T1 behave in a more complex manner. The onset of internal rotation may make T1 larger or smaller, depending on the value of τRωC. In the extreme narrowing limit, T1 increases monotonically as τG decreases, reaching a limiting value (when τG≫τR) of 4(1−3 cos2θ)−2T1R, where θ is the angle between the C–H vector and the axis of internal rotation and T1R is the value of T1 in the absence of internal rotation. However, for very slow over-all reorientation (with respect to the resonance frequency), the onset of internal rotation produces a decrease in T1 until it reaches a minimum and increases again. When τG→ 0, T1 reaches the same limiting value as in the extreme narrowing case. As expected, in the extreme narrowing limit for τR we get the full NOE of 2.988 regardless of the value of τG. For slow over-all reorientation, the onset of internal rotation first increases the NOE. As τG gets smaller, the NOE goes through a maximum and then decreases again. As τG→ 0, the NOE reaches an asymptotic value equal to that in the absence of internal rotation.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTConformation and segmental motion of native and denatured ribonuclease A in solution. Application of natural-abundance carbon-13 partially relaxed Fourier transform nuclear magnetic resonanceAdam Allerhand, David Doddrell, Victor Glushko, David W. Cochran, Ernest Wenkert, Peter J. Lawson, and Frank R. N. GurdCite this: J. Am. Chem. Soc. 1971, 93, 2, 544–546Publication Date (Print):January 1, 1971Publication History Published online1 May 2002Published inissue 1 January 1971https://pubs.acs.org/doi/10.1021/ja00731a053https://doi.org/10.1021/ja00731a053research-articleACS PublicationsRequest reuse permissionsArticle Views183Altmetric-Citations141LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-Alertsclose Get e-Alerts

Proton-decoupled Fourier transform nuclear magnetic resonance of natural abundance 13C was used to obtain spectra of ribonuclease A and its oxidized derivative at a set of pH values ranging from pH 1.40 to 6.72 at 45°. Observations on the native conformation were made at pH 4.14 and 6.55, and on the denatured conformation at pH 2.12 and below. In addition, the disrupted structure obtained by performic acid oxidation of the disulfide bonds was observed at several pH values. The loss of the native structure was associated with sharpening of the resonance bands. A tentative assignment was made for the unmodified ribonuclease A at pH 1.46 based on resonance positions of free amino acids and peptides. Although a good fit to the data was obtained, further development is required for unequivocal assignment. Partially relaxed spectra were used to obtain spin-lattice relaxation times (T1) for certain carbon types. These values are traced through several pH values for both ribonuclease A and its oxidized derivatives. Pairs of effective rotational correlation times (τeff) were derived from the T1 values of certain aliphatic resonances. By considering data from fluorescence depolarization experiments and observed line widths, it was possible to exclude one of the τeff values in the majority of cases. The results were interpreted in terms of increased segmental mobility in the denatured states, especially for α carbon nuclei that appear relatively restrained in the native state.

Abstract Proton-decoupled Fourier transform nuclear magnetic resonance of natural abundance 13C was used to obtain spectra of cyanoferrimyoglobins of harbor seal (Phoca vitulina) and sperm whale (Physeter catodon). These spectra were compared with those of hen egg white lysozyme and of bovine pancreatic ribonuclease A to show differences reflecting gross composition as well as differences tentatively attributable to more subtle conformational constraints. The cyanoferrimyoglobin spectra were much alike but not identical. Carboxymethylation of the native seal protein produced barely detectable changes at the chemical shift positions of the major adducts but no systematic alterations elsewhere in the spectrum. However, extensive modification of histidine and methionine residues by carboxymethylation of seal apomyoglobin in the presence of 8 m urea yielded a denatured product, judged by circular dichroism measurements, for which the 13C spectrum was sharp and characteristic of a denatured protein. This spectrum was used to tabulate tentative assignments. Partially relaxed spectra were used to obtain spin-lattice relaxation times (T1) for certain carbon types. These values are compared for the various myoglobin preparations mentioned above. The similarities between the native forms are striking. The denatured preparation gives many evidences of increased segmental mobility, especially for those nuclei that appear relatively restrained in the native state.