Water flip-back

Kuboniwa H, Grezesiek S, Delaglio F, Bax A. Measurement of Hn-H J couplings in calcium-free calmodulin using new 2D and 3D water-flip-back methods. J Biomol NMR 1994 4 871-878. 

Fig. 3. HNCA (a) and two implementations of HNCA-TROSY (b-c) experiments for recording intraresidual HN(/), 15N(/), 13C"(i) and sequential 1 HN(7), l5N(/), 13Ca(i — 1) correlations in 13C/15N/2H labelled proteins. Narrow and wide bars correspond to 90° and 180° flip angles, respectively, applied with phase x unless otherwise indicated. Half-ellipse denotes water selective 90° pulse to obtain water-flip-back.88,89 All 90°...

Fig. 9.1 Basic HNN COSY pulse sequence. Narrow and wide pulses correspond to flip angles of 90° and 180°, respectively, whereas low-power (water flip-back) 90° l-l pulses are illustrated as smaller narrow pulses. Delays 8 = 2.25 ms 7=15 ms (can be shorter or longer) (a = 2.5 ms fb = 0.25 ms fc= 2.25 ms fd = 0.5 ms. Unless indicated, the phase of all pulses are applied along...

The effect could be considerable for solvent-exposed parts of the backbone and could render the NOE values inaccurate. These systematic errors could be minimized by using water flip-back pulses in order to avoid saturation of H20 magnetization [11]. The NOE data are generally more susceptible to errors than Ri and R2 because (i) the NOE experiments start with the equilibrium 15N magnetization that is 10 fold lower than that of (XH) in the Hi and R2 experiments, hence relatively low sensitivity, and (ii) the NOE values are derived from only two sets of measurements, whereas R1 and R2 data are obtained from fitting multiple sets of data the latter is expected to result in a more efficient averaging of experimental errors. 

The 15N,1H shift correlation maps are most conveniently recorded with a sensitivity-enhanced HSQC sequence with incorporated water flip-back pulses for reduced saturation transfer and pulsed-field gradients for coherence selection. The pulse sequence of the experiment is shown in Fig. 14.4 A. 

Fig. 14.4 Pulse sequences used for the experiments described in this chapter. A [ N HJ-HSQC with water flip back and PFGs. The shaped pulse on the proton channel is a sine-shaped, 1.5 ms soft pulse all other pulses are hard pulses. Gradients are applied as square or sine-shaped pulses. The sign of the last gradient is reversed for anti-echo selection together with the sign of phase 6. B CPMG sequence. C bpPFGLED sequence. The delay T denotes the diffusion delay. Typically, r is set to 1 ms, T to 50-100 ms and Te to 1.2 ms.

Another commonly used technique is the water flip-back pulse, a shaped pulse designed to selectively rotate only the water magnetization by 90°, putting it back on the +z axis after a hard (nonselective) pulse has rotated all of the sample magnetization into the x-y plane. Water can be viewed as a wild and powerful bucking bronco—it must be tamed and never allowed to get out of its pen. The best place for water is on the +z axis where it will not do any harm. This is the rationale behind the flip-back pulse every time water is moved from the +z axis, use a selective pulse to put it back there. 

HOHAHA experiment with minimal water saturation was developed by Schleucher et al. (1995a) and Dhalluin et al. (1996) proposed a water-flip-back TOCSY where water-selective pulses are used to flip a maximal fraction of the water magnetization back along the +z axis at the start of the acquisition time. TOCSY experiments with excellent water suppression based on excitation sculpting (Stott et al., 1995 Hwang et al., 1995) were reported by Callihan et al. (1996). 

Figure 2 Gradient-echo-based water suppression pulse sequences, (a) WATERGATE (b) water-flip-back (c) excitation sculpting (d and e) examples of the S pulse train that is sandwiched between the gradient echo (d) water-selective inversion...

Fig. 24. Various NOESY-water suppression combinations. (A) NOESY using presaturation in the relaxation delay and mixing times, ffi) WATERGATE NOESY and (C) the water flip-back WATERGATE NOESY. ...

Fig. 26. IWo-dimensional NOESY experiment (conditions as forRg. 25), without (A) and with (B) RD amplification used as a water flip-back pulse. The spectrum in (B) has noticeably better water suppression due to the complete alignment of the water magnetization along the +z axis at the end of the detection peri. (From Abergel et al. with permission.)...

Water molecules flip back and forth as the microwave field oscillates. 

Fig. 20. A fast flip-back WATERGATE HSQC (FHSQC) that avoids water saturation and radiation damping. The WATERGATE subunit uses a 3-9-19 pulse. Details of the phase cycling, delays and gradient strengths can be found elsewhere. ...

Fig. 22. Fingerprint regions of 2D TOCSY spectra of angiotensin (0.7 mM in 90 10 HaOrHaO, pH 7.0) at 298 K acquired at 500 MHz. (A) TOCSY with presaturation, (B) basic TOCSY-WATERGATE and (C) flip-back TOCSY-WATERGATE. There is considerable bleaching of cross-peaks near the water frequency in (A). Spectrum (B) is cleaner with some cross-peaks visible at the water frequency. TTie ffip-back TOCSY-WATERGATE (C) has improved suppression with increased intensity of many of the cross-peaks. (From Fulton et al. with permission.)...

Solvent suppression. General formalism to describe the evolution of the water polarisation in the course of an NMR experiment was presented by Hiller et al The model calculations were used to optimise the efficiency of water suppression. These have demonstrated that the performance of flip-back methods improve with the increase of the longitudinal relaxation rate of water protons. To this aim relaxation agents that selectively enhance water relaxation can be successfully used. 

Otters are extremely playful. Anyone who visits a zoo can see otters sliding down embankments and flipping about in the water. Otters typically twist and turn as they swim, sometimes on their back and sometimes on their side. Having seemingly unlimited energy, otters often slide down muddy banks over and over again with no apparent motive other than sheer enjoyment. 

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