Probing chemical shifts of invisible states of proteins with relaxation dispersion NMR spectroscopy: how well can we do?
Carr-Purcell-Meiboom-Gill relaxation dispersion NMR spectroscopy has evolved into a powerful approach for the study of low populated, invisible conformations of biological molecules. One of the powerful features of the experiment is that chemical shift differences between the exchanging conformers can be obtained, providing structural information about invisible excited states. Through the development of new labeling approaches and NMR experiments it is now possible to measure backbone 13C(alpha) and 13CO relaxation dispersion profiles in proteins without complications from 13C-13C couplings. Such measurements are presented here, along with those that probe exchange using 15N and 1HN nuclei. A key experimental design has been the choice of an exchanging system where excited-state chemical shifts were known from independent measurement. Thus it is possible to evaluate quantitatively the accuracy of chemical shift differences obtained in dispersion experiments and to establish that in general very accurate values can be obtained. The experimental work is supplemented by computations that suggest that similarly accurate shifts can be measured in many cases for systems with exchange rates and populations that fall within the range of those that can be quantified by relaxation dispersion. The accuracy of the extracted chemical shifts opens up the possibility of obtaining quantitative structural information of invisible states of the sort that is now available from chemical shifts recorded on ground states of proteins.
Hansen
D Flemming
DF
Department of Medical Genetics, University of Toronto, Toronto, Ontario, Canada, M5S 1A8.
Vallurupalli
Pramodh
P
Lundström
Patrik
P
Neudecker
Philipp
P
Kay
Lewis E
LE
D003198:Computer Simulation; D009682:Magnetic Resonance Spectroscopy / Q000379:methods* / Q000592:standards*; D008840:Microfilament Proteins / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry*; D012015:Reference Standards; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*
C062612:ABP1 protein, S cerevisiae; D008840:Microfilament Proteins; D029701:Saccharomyces cerevisiae Proteins; C491504:ARK1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases