Facile measurement of polypeptide JHNHα coupling constants from HMQC-J spectra

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A method, based on linewidth measurements, is described which permits the rapid and facile determination of JHNHα coupling constants from 15N labeled proteins. Using appropriately processed HMQC-J data, we have found that a simple linear relationship exists between the half-height linewidth (Δν1/2) of 15N–1H cross peaks and their corresponding JHNHα coupling constants. Tests indicate that this technique permits the accurate measurement of up to 100 JHNHα coupling constants in less than 30 min. Furthermore, the Jα measurements can be done manually – without the need of any computer-based curve-fitting or minimization. Comparisons between JHNHα values predicted from high resolution X-ray structures and those determined using this technique indicate that the method is both accurate and precise (correlation coefficient = 0.90, rmsd = 0.75 Hz).

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