TY - JOUR
T1 - DNP-Assisted NMR Investigation of Proteins at Endogenous Levels in Cellular Milieu
AU - Costello, Whitney N.
AU - Xiao, Yiling
AU - Frederick, Kendra K.
PY - 2018/1/1
Y1 - 2018/1/1
N2 - Structural investigations of biomolecules are typically confined to in vitro systems under extremely limited conditions. These investigations yield invaluable insights, but such experiments cannot capture important structural features imposed by cellular environments. Structural studies of proteins in their native contexts are not only possible using state-of-the-art sensitivity-enhanced (dynamic nuclear polarization, DNP) solid-state nuclear magnetic resonance (NMR) techniques, but these studies also demonstrate that the cellular context can and does have a dramatic influence on protein structure. In this chapter, we describe methods to prepare samples of isotopically labeled proteins at endogenous levels in cellular contexts alongside quality control methods to ensure that such samples accurately model important features of the cellular environment.
AB - Structural investigations of biomolecules are typically confined to in vitro systems under extremely limited conditions. These investigations yield invaluable insights, but such experiments cannot capture important structural features imposed by cellular environments. Structural studies of proteins in their native contexts are not only possible using state-of-the-art sensitivity-enhanced (dynamic nuclear polarization, DNP) solid-state nuclear magnetic resonance (NMR) techniques, but these studies also demonstrate that the cellular context can and does have a dramatic influence on protein structure. In this chapter, we describe methods to prepare samples of isotopically labeled proteins at endogenous levels in cellular contexts alongside quality control methods to ensure that such samples accurately model important features of the cellular environment.
KW - Amyloids
KW - Dynamic nuclear polarization
KW - In situ structural biology
KW - In-cell NMR
KW - NMR
KW - Prions
KW - Sup35
UR - http://www.scopus.com/inward/record.url?scp=85053714787&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85053714787&partnerID=8YFLogxK
U2 - 10.1016/bs.mie.2018.08.023
DO - 10.1016/bs.mie.2018.08.023
M3 - Article
C2 - 30638534
AN - SCOPUS:85053714787
SN - 0076-6879
JO - Methods in Enzymology
JF - Methods in Enzymology
ER -