期刊
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
卷 140, 期 6, 页码 2115-2126出版社
AMER CHEMICAL SOC
DOI: 10.1021/jacs.7b09576
关键词
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资金
- Canadian Institutes of Health Research
- Natural Sciences and Engineering Research Council of Canada
R-1 rho relaxation dispersion experiments are increasingly used in studies of protein dynamics on the micro- to millisecond time scale. Traditional R-1 rho relaxation dispersion approaches are typically predicated on changes in chemical shifts between corresponding probe spins, Delta omega(GE), in the interconverting states. Here, we present a new application of off-resonance N-15 R-1 rho relaxation dispersion that enables the quantification of slow exchange processes even in the limit where Delta omega(GE) = 0 so long as the spins in the exchanging states have different intrinsic transverse relaxation rates (Delta R-2 = R-2,R-E - R-2,R-G not equal 0). In this limit, the dispersion profiles become inverted relative to those measured in the case where Delta omega(GE) not equal 0, Delta R-2 = 0. The theoretical background to understand this effect is presented, along with a simplified exchange matrix that is valid in the limits that are germane here. An application to the study of the dynamics of the germ granule protein Ddx4 in a highly concentrated phase-separated state is described. Notably, exchange-based dispersion profiles can be obtained despite the fact that Delta omega(GE) approximate to 0 and Delta R-2 is small, similar to 20-30 s(-1). Our results are consistent with the formation of a significantly populated excited conformational state that displays increased contacts between adjacent protein molecules relative to the major conformer in solution, leading to a decrease in overall motion of the protein backbone. A complete set of exchange parameters is obtained from analysis of a single set of N-15 off-resonance R-1 rho measurements recorded at a single static magnetic field and with a single spin-lock radio frequency field strength. This new approach holds promise for studies of weakly interacting systems, especially those involving intrinsically disordered proteins that form phase-separated organelles, where little change to chemical shifts between interconverting states would be expected, but where finite Delta R-2 values are observed.
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