Heat Management Strategies for Solid-state NMR of Functional Proteins
 

Heat Management Strategies for Solid-state NMR of Functional Proteins


Publication year: 2012
Source:Journal of Magnetic Resonance</br>
Daniel J. Fowler, Michael J. Harris, Lynmarie K. Thompson</br>
Modern solid-state NMR methods can acquire high-resolution protein spectra for structure determination. However, these methods use rapid sample spinning and intense decoupling fields that can heat and denature the protein being studied. Here we present a strategy to avoid destroying valuable samples. We advocate first creating a sacrificial sample, which contains unlabeled protein (or no protein) in buffer conditions similar to the intended sample. This sample is then doped with the chemical shift thermometer Sm2Sn2O7. We introduce a pulse scheme called TCUP (for Temperature Calibration Under Pulseload) that can characterize the heating of this sacrificial sample rapidly, under a variety of experimental conditions, and with high temporal resolution. Sample heating is discussed with respect to different instrumental variables such as spinning speed, decoupling strength and duration, and cooling gas flow rate. The effects of different sample preparation variables are also discussed, including ionic strength, the inclusion of cryoprotectants, and the physical state of the sample (i.e. liquid, solid, or slurry). Lastly, we discuss probe detuning as a measure of sample thawing that does not require retuning the probe or using chemical shift thermometer compounds. Use of detuning tests and chemical shift thermometers with representative sample conditions makes it possible to maximize the efficiency of the NMR experiment while retaining a functional sample.
Graphical abstract

http://origin-ars.els-cdn.com/conten...002352-fx1.jpg Graphical abstract Highlights

? Heating tests on a sacrificial sample enable efficient NMR of functional proteins. ? Sm2Sn2O7 chemical shift thermometer doesn’t alter freezing point or ionic strength. ? Complete freezing of high salt samples minimizes heating. ? Simple tests for thawing during pulsing are described. ? Increased cooling efficiency is an untapped method for effective heat management.</br>
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Source: Journal of Magnetic Resonance