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J Biomol NMR. 1994 Nov;4(6):871-8.

Measurement of HN-H alpha J couplings in calcium-free calmodulin using new 2D and 3D water-flip-back methods.

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Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520.


Two new methods are described for the measurement of three-bond JHNH alpha couplings in proteins isotopically enriched with 15N. Both methods leave the water magnetization in an unsaturated state, parallel to the z-axis, and therefore offer significant enhancements in sensitivity for rapidly exchanging backbone amide protons. The J couplings can be measured either from a set of constant-time 2D 1H-15N HMQC spectra, which are modulated in intensity by JHNH alpha, or from a water-flip-back version of the 3D HNHA experiment. The method is demonstrated for a sample of calcium-free calmodulin. Residues Lys75-Asp80 have JHNH alpha values in the 6-7 Hz range, suggesting that a break in the 'central helix' occurs at the same position as previously observed in solution NMR studies of Ca(2+)-ligated calmodulin.

[Indexed for MEDLINE]

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