The relevance of J cross-peaks in two-dimensional NOE experiments of macromolecules

Abstract

Two-dimensional (2D) exchange spectroscopy (I, 2) is on the point of becoming a major technique for the elucidation of the spatial structure of biological macromolecules through the nuclear Overhauser effect (NOE) (3-8) and for the investigation of chemical exchange processes (9, IO). In particular, two-dimensional NOE spectroscopy has already been well established as a powerful tool for investigating proteins in solution (3, 4, 6-8). The nuclear Overhauser effect and chemical exchange cause an incoherent exchange of magnetization between different sites, either by cross-relaxation or by an actual exchange of nuclei, respectively. In 2D spectroscopy these processes are traced out by “frequency-labeling” the various magnetization components before exchange and by sorting out the labeled components after exchange (1). In addition to the cross-peaks caused by this incoherent exchange, so-called J cross-peaks may arise as a consequence of coherent transfer of magnetization between J-coupled resonances (I I, 12). Although the J cross-peaks contain valuable information on the coupling network, which is exploited in 2D-correlated spectroscopy (23-15) they are undesired in exchange spectra. They may be misinterpreted as additional exchange cross-peaks or may falsify the measured intensities in the case of coincidence with exchange cross-peaks. Suitable techniques for their suppression have been described recently (I 1, 12). In this communication, we would like to quantitatively evaluate the relevance of J cross-peaks in NOESY spectra of proteins by means of experimental measurements and by simple model calculations. Zero-quantum coherence precessing during the mixing period of duration T, is primarily responsible for the occurrence of J cross-peaks. Double-quantum coherence may also contribute. However, by proper phase-cycling (II), it is possible to eliminate J cross-peak contributions originating from double-quantum coherence. The interference effects from transverse magnetization present during the mixing time are usually suppressed by a simple phase alternation technique. Higher-order multiple-quantum coherence is seldom of practical significance for the creation of J cross-peaks.