Skip to main content
. 1998 Nov 24;95(24):14147–14151. doi: 10.1073/pnas.95.24.14147

Figure 2.

Figure 2

Experimental scheme for the 2D hJNN-correlation-[15N,1H]-TROSY experiment used to correlate the chemical shifts of pairs of hydrogen bond-related 15N spins and to measure hJHN. On the lines marked 1H, 15N, and 13C, narrow and wide bars stand for nonselective 90° and 180° radio frequency pulses, respectively, with the carrier frequencies at 12, 190, and 145 ppm, respectively. The delays are τ1 = 2.7 ms, τ2 = 18 ms, and τ3 = τ2 − τ1. The line marked PFG indicates the pulsed magnetic field gradients applied along the z axis: G1, amplitude 30 G/cm, duration 1 ms; G2, 42 G/cm, 1 ms; G3, 45 G/cm, 1 ms; G4, 40 G/cm, 1 ms; and G5, 48 G/cm, 1 ms. Two data sets, I and II, are measured in the interleaved manner with the following phase cycling schemes: (I), φ1 = {−x, x}; φ2 = {2(−x),2x}; ψ1 = {4y,4(−y)}; ψ2(receiver) = {y,−y,−y, y,−y, y,y,−y}; (II), φ1 = {−y, y}; φ2 = {2(−y),2y}; ψ1 = {4(−x),4x}; ψ2(receiver) = {−x, x,x,−x, x,−x,−x, x}; x on all other pulses. For both data sets, quadrature detection in the t1 dimension is obtained by using States-TPPI (38) by simultaneously incrementing each of the phases φ2 and ψ2 by 90°. The transfer of 15N magnetization to 1H for detection of single transition-to-single transition polarization transfer (5) was used. Fourier transformation of the sum and difference of the two data sets results in two 2D hJNN- correlation-[15N,1H]-TROSY spectra in which the relative cross-peak positions are determined in E.COSY manner by the chemical shifts and the scalar-coupling constants (27–30). For example, both the direct and the relayed correlation cross-peaks that appear in the two spectra are shifted by 1JHN along ω2(1H). Along ω1(15N), the direct peaks also are shifted by 1JHN, but the separation of the relayed cross-peaks is determined exclusively by hJHN. To enhance the signals of rapidly exchanging imino protons, water saturation is minimized by keeping the water magnetization along +z during the entire experiment, using the water-selective 90° pulses indicated by curved shapes on the line 1H.