Direct determination of the heteronuclear T-1/T-2 ratio by off-resonance steady-state magnetization measurement: Investigation of the possible application to fast exchange characterization of N-15-labeled proteins

The N-15 steady-state magnetization in the presence of off-resonance rf irr adiation is an analytical function of the T-1/T-2 ratio and of the angle be tween the N-15 effective field axis and the static magnetic field direction . This relation holds whatever the relaxation mechanisms due to motions on the nanosecond time scale, and the size of the spin system. If motions on t he micro- to millisecond time scale are present (fast exchange), the same o bservable depends also on their spectral density at the frequency of the ef fective field. The cross-peak intensity in each 2D N-15-H-1 correlation map is directly related to the dynamic parameters, so that the characterizatio n of fast exchange phenomena by this method is in principle less time-consu ming than the separate measurement of self-relaxation rates. The theory of this approach is described. Its practical validity is experimentally evalua ted on a N-15-labeled 61 amino acid neurotoxin. It turns out that existing equipments lead to non-negligible biases. Their consequences for the accura cy attainable, at present, by this method are investigated in detail.