TmDOTP : An NMR- based Thermometer for Magic Angle Spinning NMR Experiments

Abstract

11Solid state NMR is a powerful tool to probe membrane protein structure and motions in native 12 lipid structures. Sample heating, caused by magic angle spinning and radio frequency irradiation 13 in solid state NMR, produces uncertainties in sample temperature and thermal broadening caused 14 by temperature distributions, which can also lead to sample deterioration. To measure the sample 15 temperature in real time, and to quantify thermal gradients and their dependence on radio 16 frequency irradiation or spinning frequency, we use the chemical shift thermometer TmDOTP, a 17 lanthanide complex. Compared to other NMR thermometers (e.g., the proton NMR signal of 18 water), the proton spectrum of TmDOTP exhibits higher thermal sensitivity and resolution. In 19 addition, the H6 proton in TmDOTP has a large chemical shift (-175 ppm at 275 K) and is well 20 resolved from the rest of the proton spectrum. We identified two populations of TmDOTP, with 21 differing temperatures and dependency on the radio frequency irradiation power, within 22 proteoliposome samples. We interpret these populations as arising from the supernatant and the 23 pellet, which is sedimented from the sample spinning. Our results indicate that TmDOTP is an 24 excellent internal standard for monitoring temperatures of biophysically relevant samples 25 without distorting their properties. 26 27Introduction 28Magic angle spinning (MAS) solid-state nuclear magnetic resonance (SSNMR) is a powerful 29 technique for studying biomolecules 1 , including: protein assemblies in near native conditions 2 1, 30 membrane proteins 3-5 and amyloid fibrils [6][7][8] . SSNMR provides rich information on protein 31 molecular structure and motions. Restricted global molecular motions in solids allow for the 32 retention of dipolar couples, which enables direct measurement of distances and local orientations. 33Many of the most exciting developments in SSNMR however, involve pulse sequences with long 34 and strong radio frequency (RF) irradiation elements; Sample heating from magic angle spinning 35and RF irradiation has been cause for concern 9,10 Elevated and uncalibrated temperatures within 36 the sample complicates the interpretation of dynamics and other properties. Moreover, heating 37 gradients within the MAS rotor may contribute to peak broadening. 38 39Sample heating originates in part from friction between bearing gas and the rotor during MAS 10,11 . 40 Heat is also generated from RF irradiation during high power decoupling due to inductive 41 dielectric heating on conductive or dipolar samples 9,12,13 . The application of high power oscillating 42 electric field causes free charges and permanent electric dipoles to move, generating kinetic 43 energy , which dissipates in the surrounding sample as heat 14 . The absorption of RF energy is 44 maximized when ωτ = 1, where ω represents the frequency of oscillating field and τ is the 45 characteristic relaxation time of the molecule 14,15 . RF heating is of particular concern in SSNMR 46