AN EVALUATION OF LEAST-SQUARES FITS TO COSY SPECTRA AS A MEANS OF ESTIMATING PROTON-PROTON COUPLING-CONSTANTS .2. APPLICATIONS TO POLYPEPTIDES

A new computational method for simultaneously estimating all the proto n-proton coupling constants in a molecule from COSY spectra [Yang, J.- X. and Havel, T.F. (1994) J. Biomol. NMR, 4, 807-826] is applied to ex perimental data from two polypeptides. The first of these is a cyclic hexapeptide denoted as VDA -D-Ala(1)-Phe(2)-Trp(3)-Lys(Z)(4)-Val(5)-Ph e(6)-), in deuterated DMSO, while the second is a 39-residue protein, called decorsin, in aqueous solution. The effect of different data pro cessing strategies and different initial parameter values on the accur acy of the coupling constants was explored. In the case of VDA, most o f the coupling constants did not depend strongly on the initial values chosen for the optimization or on how the data were processed. This, together with our previous experience using simulated data, implies st rongly that these values are accurate estimates of the coupling consta nts. They also differ by an average of only 0.36 Hz from the values of the 14 coupling constants that could be measured independently by est ablished methods. In the case of decorsin, many of the coupling consta nts exhibited a moderate dependence on their initial values and a stro ng dependence on how the data were processed. With the most successful data processing strategy, the amide-alpha coupling constants differed by an average of 1.11 Hz from the 21 values that could be measured by established methods, while two thirds of the three-bond coupling cons tants fell within 1.0 Hz of the ranges obtained by applying the Karplu s relation to an independently computed ensemble of distance geometry structures. The averages of the coupling constants over multiple optim izations using random initial values were computed in order to obtain the best possible estimates of the coupling constants. Most clearly in correct averages can be identified by large standard deviations in the coupling constants or the associated line widths and chemical shifts, and can be explained by strong coupling and/or overlap with the water signal, the diagonal peaks or other cross peaks.