skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Single-Molecule Protein Conformational Dynamics in Cell Signaling

Abstract

We have demonstrated the application of single-molecule imaging and ultrafast spectroscopy to probe protein conformational dynamics in solution and in lipid bilayers. Dynamic protein-protein interactions involve significant conformational motions that initiate chain reactions leading to specific cellular responses. We have carried out a single molecule study of dynamic protein-protein interactions in a GTPase intracellular signaling protein Cdc42 in complex with a downstream effector protein, WASP. We were able to probe hydrophobic interactions significant to Cdc42/WASP recognition. Single molecule fluorescence intensity and polarization measurements have revealed the dynamic and inhomogeneous nature of protein-protein interactions within the Cdc42/WASP complex that is characterized by structured distributions of conformational fluctuation rates. Conducting a single-molecule fluorescence anisotropy study of calmodulin (CaM), a regulatory protein for calcium-dependent cell signaling, we were able to probe CaM conformational dynamics at a wide time scale. In this study, CaM contains a site-specifically inserted tetra-cysteine motif that reacted with FlAsH, a biarsenic fluorescein derivative that can be rotationally locked to the host protein. The study provided direct characterization of the nanosecond motions of CaM tethered to a biologically compatible surface under physiological buffer solution. The unique technical approaches are applicable of studying single-molecule dynamics of protein conformational motions and protein-proteinmore » interactions at a wide time range without the signal convolution of probe-dye molecule motions« less

Authors:
Publication Date:
Research Org.:
Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
Sponsoring Org.:
USDOE
OSTI Identifier:
15020318
Report Number(s):
PNNL-SA-44493
6506; KC0301020; TRN: US200519%%365
DOE Contract Number:  
AC05-76RL01830
Resource Type:
Conference
Resource Relation:
Conference: American Chemical Society, 228(Pt. 2):Phys 711; American Chemical Society,Washington,,United States.
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ANISOTROPY; BUFFERS; CALMODULIN; CHAIN REACTIONS; FLUCTUATIONS; FLUORESCEIN; FLUORESCENCE; LIPIDS; POLARIZATION; PROBES; PROTEINS; SPECTROSCOPY; environmental molecular sciences laboratory

Citation Formats

Lu, H PETER. Single-Molecule Protein Conformational Dynamics in Cell Signaling. United States: N. p., 2004. Web.
Lu, H PETER. Single-Molecule Protein Conformational Dynamics in Cell Signaling. United States.
Lu, H PETER. Sun . "Single-Molecule Protein Conformational Dynamics in Cell Signaling". United States.
@article{osti_15020318,
title = {Single-Molecule Protein Conformational Dynamics in Cell Signaling},
author = {Lu, H PETER.},
abstractNote = {We have demonstrated the application of single-molecule imaging and ultrafast spectroscopy to probe protein conformational dynamics in solution and in lipid bilayers. Dynamic protein-protein interactions involve significant conformational motions that initiate chain reactions leading to specific cellular responses. We have carried out a single molecule study of dynamic protein-protein interactions in a GTPase intracellular signaling protein Cdc42 in complex with a downstream effector protein, WASP. We were able to probe hydrophobic interactions significant to Cdc42/WASP recognition. Single molecule fluorescence intensity and polarization measurements have revealed the dynamic and inhomogeneous nature of protein-protein interactions within the Cdc42/WASP complex that is characterized by structured distributions of conformational fluctuation rates. Conducting a single-molecule fluorescence anisotropy study of calmodulin (CaM), a regulatory protein for calcium-dependent cell signaling, we were able to probe CaM conformational dynamics at a wide time scale. In this study, CaM contains a site-specifically inserted tetra-cysteine motif that reacted with FlAsH, a biarsenic fluorescein derivative that can be rotationally locked to the host protein. The study provided direct characterization of the nanosecond motions of CaM tethered to a biologically compatible surface under physiological buffer solution. The unique technical approaches are applicable of studying single-molecule dynamics of protein conformational motions and protein-protein interactions at a wide time range without the signal convolution of probe-dye molecule motions},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2004},
month = {8}
}

Conference:
Other availability
Please see Document Availability for additional information on obtaining the full-text document. Library patrons may search WorldCat to identify libraries that hold this conference proceeding.

Save / Share: