TY - JOUR
T1 - Sensitivity-Enhanced NMR Reveals Alterations in Protein Structure by Cellular Milieus
AU - Frederick, Kendra K.
AU - Michaelis, Vladimir K.
AU - Corzilius, Björn
AU - Ong, Ta Chung
AU - Jacavone, Angela C.
AU - Griffin, Robert G.
AU - Lindquist, Susan
N1 - Funding Information:
We thank the members of the S.L. and R.G.G. groups for valuable discussions and comments during the course of this research. S.L. is an investigator of the Howard Hughes Medical Institute. K.K.F. was supported by the Life Science Research Foundation as an HHMI fellow. V.K.M. is grateful to the Natural Sciences and Engineering Research Council of Canada and the Government of Canada for a Banting postdoctoral fellowship. B.C. was supported by the Deutsche Forschungsgemeinschaft (research fellowship CO 802/1-1). This work was funded by grants from the G. Harold and Leila Y. Mathers Foundation (S.L.) and by NIH grants GM-025874 to S.L. and EB-003151, EB-002804, and EB-002026 to R.G.G.
Publisher Copyright:
© 2015 Elsevier Inc.
PY - 2015/10/22
Y1 - 2015/10/22
N2 - Summary Biological processes occur in complex environments containing a myriad of potential interactors. Unfortunately, limitations on the sensitivity of biophysical techniques normally restrict structural investigations to purified systems, at concentrations that are orders of magnitude above endogenous levels. Dynamic nuclear polarization (DNP) can dramatically enhance the sensitivity of nuclear magnetic resonance (NMR) spectroscopy and enable structural studies in biologically complex environments. Here, we applied DNP NMR to investigate the structure of a protein containing both an environmentally sensitive folding pathway and an intrinsically disordered region, the yeast prion protein Sup35. We added an exogenously prepared isotopically labeled protein to deuterated lysates, rendering the biological environment "invisible" and enabling highly efficient polarization transfer for DNP. In this environment, structural changes occurred in a region known to influence biological activity but intrinsically disordered in purified samples. Thus, DNP makes structural studies of proteins at endogenous levels in biological contexts possible, and such contexts can influence protein structure.
AB - Summary Biological processes occur in complex environments containing a myriad of potential interactors. Unfortunately, limitations on the sensitivity of biophysical techniques normally restrict structural investigations to purified systems, at concentrations that are orders of magnitude above endogenous levels. Dynamic nuclear polarization (DNP) can dramatically enhance the sensitivity of nuclear magnetic resonance (NMR) spectroscopy and enable structural studies in biologically complex environments. Here, we applied DNP NMR to investigate the structure of a protein containing both an environmentally sensitive folding pathway and an intrinsically disordered region, the yeast prion protein Sup35. We added an exogenously prepared isotopically labeled protein to deuterated lysates, rendering the biological environment "invisible" and enabling highly efficient polarization transfer for DNP. In this environment, structural changes occurred in a region known to influence biological activity but intrinsically disordered in purified samples. Thus, DNP makes structural studies of proteins at endogenous levels in biological contexts possible, and such contexts can influence protein structure.
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U2 - 10.1016/j.cell.2015.09.024
DO - 10.1016/j.cell.2015.09.024
M3 - Article
C2 - 26456111
AN - SCOPUS:84948961356
VL - 163
SP - 620
EP - 628
JO - Cell
JF - Cell
SN - 0092-8674
IS - 3
ER -