Fast multi-dimensional NMR of proteins.

Three-dimensional HNCO and HNCA subspectra from a small protein (agitoxin, 4 kDa, enriched in carbon-13 and nitrogen-15), have been obtained by direct frequency-domain excitation of selected carbon and nitrogen sites. This new technique applies an array of several simultaneous soft radiofrequency spin-inversion pulses, encoded (on or off) according to nested Hadamard matrices, and the resulting responses are extracted by reference to the same matrices. This not only simplifies the spectra but, by avoiding extensive sampling in the traditional evolution dimensions, affords a speed advantage of more than two orders of magnitude.