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Water proton spin saturation affects measured protein backboneN spin relaxation rates


Publication year: 2011
Source: Journal of Magnetic Resonance, Available online 1 October 2011

Kang*Chen, Nico*Tjandra

Protein backboneN NMR spin relaxation rates are useful in characterizing the protein dynamics and structures. To observe the protein nuclear-spin resonances a pulse sequence has to include a water suppression scheme. There are two commonly employed methods, saturating or dephasing the water spins with pulse field gradients and keeping them unperturbed with flip-back pulses. Here different water suppression methods were incorporated into pulse sequences to measureN longitudinalT1and transversal rotating-frameT1?spin relaxation. Unexpectedly theNT1relaxation time constants varied significantly with the choice of water suppression method. For a 25-kDaE.coli. glutamine binding protein (GlnBP) theT1values acquired with the pulse sequence containing a water dephasing gradient are on average 20% longer than the ones obtained using a pulse sequence containing the water flip-back pulse. In contrast the twoT1 ?data sets are correlated without an apparent offset. The averageT1difference was reduced to 12% when the experimental recycle delay was doubled, while the averageT1values from the flip-back measurements were nearly unchanged. Analysis of spectral signal to noise ratios (s/n) showed the apparent slowerN relaxation obtained with the water dephasing experiment originated from the differences inHNrecovery for each relaxation time point. This in turn offset signal reduction fromN relaxation decay. The artifact becomes noticeable when the measuredN relaxation time constant is comparable to recycle delay, e.g., theNT1of medium to large proteins. TheN relaxation rates measured with either water suppression schemes yield reasonable fits to the structure. However, data from the saturated scheme results in significantly lower Model-Free order parameters ( = 0.81) than the non-saturated ones ( = 0.88), indicating such order parameters may be previously underestimated.

Graphical abstract



Highlights

? Different water suppression methods were adopted to measure NMR relaxation. ? SlowerNT1rates were identified when the water was saturated in the experiment. ? The cause for the slowerT1relaxation is the non-uniformHNrecovery. ? SlowerT1relaxation leads to apparent lower Model-Free order parameters.



Source: Journal of Magnetic Resonance