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Role of Protein Stabilizers on the Conformation of the Unfolded State of Cytochrome c and Its Early Folding Kinetics

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Title Role of Protein Stabilizers on the Conformation of the
Unfolded State of Cytochrome c and Its Early Folding Kinetics
 
Creator Haldar, Shubhasis
Mitra, Samaresh
Chattopadhyay, Krishnananda
 
Subject Structural Biology & Bioinformatics
 
Description An insight into the conformation and dynamics of unfolded
and early intermediate states of a protein is essential to understand
the mechanism of its aggregation and to design potent
inhibitor molecules. Fluorescence correlation spectroscopy has
been used to study the effects of several model protein stabilizers
on the conformation of the unfolded state and early folding
dynamics of tetramethyl rhodamine-labeled cytochrome c from
Saccharomyces cerevisiae at single molecular resolution. Special
attention has been given to arginine, which is a widely used stabilizer
for improving refolding yield of different proteins. The
value of the hydrodynamic radius (rH) obtained by analyzing the
intensity fluctuations of the diffusing molecules has been found
to increase in a two-state manner as the protein is unfolded by
urea. The results further show that the presence of arginine and
other protein stabilizers favors a relatively structured conformation
of the unfolded states (rH of 29A˚ ) over an extended one
(rH of 40A˚ ), which forms in their absence. Also, the time constant
of a kinetic component (�R) of about 30 �s has been
observed by analyzing the correlation functions, which represents
formation of a collapsed state. This time constant varies
with urea concentration representing an inverted Chevron plot
that shows a roll-over behavior in the absence of arginine. To the
best of our knowledge, this is one of the first applications of
fluorescence correlation spectroscopy to study direct folding
kinetics of a protein.
 
Publisher American Society for Biochemistry and Molecular Biology
 
Date 2010
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/71/1/JOURNAL_OF_BIOLOGICAL_CHEMISTRY_285(33)25314%2D25323;2010[46].pdf
Haldar, Shubhasis and Mitra, Samaresh and Chattopadhyay, Krishnananda (2010) Role of Protein Stabilizers on the Conformation of the Unfolded State of Cytochrome c and Its Early Folding Kinetics. The Journal of Biological Chemistry, 285 (33). pp. 25314-25323.
 
Relation http://dx.doi.org/10.1074/jbc.M110.116673
http://www.eprints.iicb.res.in/71/