TY - JOUR
T1 - Quantifying internal friction in unfolded and intrinsically disordered proteins with single-molecule spectroscopy
AU - Soranno, Andrea
AU - Buchli, Brigitte
AU - Nettels, Daniel
AU - Cheng, Ryan R.
AU - Müller-Späth, Sonja
AU - Pfeil, Shawn H.
AU - Hoffmann, Armin
AU - Lipman, Everett A.
AU - Makarov, Dmitrii E.
AU - Schuler, Benjamin
PY - 2012/10/30
Y1 - 2012/10/30
N2 - Internal friction, which reflects the "roughness" of the energy landscape, plays an important role for proteins by modulating the dynamics of their folding and other conformational changes. However, the experimental quantification of internal friction and its contribution to folding dynamics has remained challenging. Here we use the combination of single-molecule Förster resonance energy transfer, nanosecond fluorescence correlation spectroscopy, and microfluidic mixing to determine the reconfiguration times of unfolded proteins and investigate the mechanisms of internal friction contributing to their dynamics. Using concepts from polymer dynamics, we determine internal friction with three complementary, largely independent, and consistent approaches as an additive contribution to the reconfiguration time of the unfolded state. We find that the magnitude of internal friction correlates with the compactness of the unfolded protein: its contribution dominates the reconfiguration time of approximately 100 ns of the compact unfolded state of a small cold shock protein under native conditions, but decreases for more expanded chains, and approaches zero both at high denaturant concentrations and in intrinsically disordered proteins that are expanded due to intramolecular charge repulsion. Our results suggest that internal friction in the unfolded state will be particularly relevant for the kinetics of proteins that fold in the microsecond range or faster. The low internal friction in expanded intrinsically disordered proteins may have implications for the dynamics of their interactions with cellular binding partners.
AB - Internal friction, which reflects the "roughness" of the energy landscape, plays an important role for proteins by modulating the dynamics of their folding and other conformational changes. However, the experimental quantification of internal friction and its contribution to folding dynamics has remained challenging. Here we use the combination of single-molecule Förster resonance energy transfer, nanosecond fluorescence correlation spectroscopy, and microfluidic mixing to determine the reconfiguration times of unfolded proteins and investigate the mechanisms of internal friction contributing to their dynamics. Using concepts from polymer dynamics, we determine internal friction with three complementary, largely independent, and consistent approaches as an additive contribution to the reconfiguration time of the unfolded state. We find that the magnitude of internal friction correlates with the compactness of the unfolded protein: its contribution dominates the reconfiguration time of approximately 100 ns of the compact unfolded state of a small cold shock protein under native conditions, but decreases for more expanded chains, and approaches zero both at high denaturant concentrations and in intrinsically disordered proteins that are expanded due to intramolecular charge repulsion. Our results suggest that internal friction in the unfolded state will be particularly relevant for the kinetics of proteins that fold in the microsecond range or faster. The low internal friction in expanded intrinsically disordered proteins may have implications for the dynamics of their interactions with cellular binding partners.
KW - Energetic roughness
KW - Kramers theory
KW - Protein folding
KW - Rouse model
KW - Single-molecule FRET
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UR - http://www.scopus.com/inward/citedby.url?scp=84862076708&partnerID=8YFLogxK
U2 - 10.1073/pnas.1117368109
DO - 10.1073/pnas.1117368109
M3 - Article
C2 - 22492978
AN - SCOPUS:84862076708
SN - 0027-8424
VL - 109
SP - 17800
EP - 17806
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 44
ER -