RRC ID 68742
Author Oura M, Yamamoto J, Ishikawa H, Mikuni S, Fukushima R, Kinjo M.
Title Polarization-dependent fluorescence correlation spectroscopy for studying structural properties of proteins in living cell.
Journal Sci Rep
Abstract Rotational diffusion measurement is predicted as an important method in cell biology because the rotational properties directly reflect molecular interactions and environment in the cell. To prove this concept, polarization-dependent fluorescence correlation spectroscopy (pol-FCS) measurements of purified fluorescent proteins were conducted in viscous solution. With the comparison between the translational and rotational diffusion coefficients obtained from pol-FCS measurements, the hydrodynamic radius of an enhanced green fluorescent protein (EGFP) was estimated as a control measurement. The orientation of oligomer EGFP in living cells was also estimated by pol-FCS and compared with Monte Carlo simulations. The results of this pol-FCS experiment indicate that this method allows an estimation of the molecular orientation using the characteristics of rotational diffusion. Further, it can be applied to analyze the degree of molecular orientation and multimerization or detection of tiny aggregation of aggregate-prone proteins.
Volume 6
Pages 31091
Published 2016-8-4
DOI 10.1038/srep31091
PII srep31091
PMID 27489044
PMC PMC4973283
MeSH Animals COS Cells Chlorocebus aethiops Fluorescence Polarization Green Fluorescent Proteins / chemistry* Hydrodynamics Molecular Dynamics Simulation Monte Carlo Method Protein Aggregates Protein Structure, Quaternary Recombinant Proteins / chemistry Spectrometry, Fluorescence / methods*
IF 3.998
DNA material FL-linked EGFP dimer (RDB19236) FL-linked EGFP trimer (RDB19237) FL-linked EGFP tetramer (RDB19238) FL-linked EGFP pentamer (RDB19239)