Two-dimensional IR correlation spectroscopy : sequential events in the unfolding process of the λ Cro-V55C repressor protein

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DOIResolve DOI: http://doi.org/10.1073/pnas.96.23.13153
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TypeArticle
Journal titleProceedings of the National Academy of Sciences of the United States of America
ISSN1091-6490
Volume96
Issue23
Pages1315313158; # of pages: 6
AbstractA question often posed in protein folding/unfolding studies is whether the process is fully cooperative or whether it contains sequential elements. To address this question, one needs tools capable of resolving different events. It seems that, at least in certain cases, two-dimensional (2D) IR correlation spectroscopy can provide answers to this question. To illustrate this point, we have turned to the Cro-V55C dimer of the λ Cro repressor, a protein known to undergo thermal unfolding in two discrete steps through a stable equilibrium intermediate. The secondary structure of this intermediate is compatible with that of a partially unfolded protein and involves a reorganization of the N terminus, whereas the antiparallel β-ribbon formed by the C-terminal part of each subunit remains largely intact. To establish whether the unfolding process involves sequential events, we have performed a 2D correlation analysis of IR spectra recorded over the temperature range of 20–95°C. The 2D IR correlation analysis indeed provides evidence for a sequential formation of the stable intermediate, which is created in three (closely related) steps. A first step entails the unfolding of the short N-terminal β-strand, followed by the unfolding of the α-helices in a second step, and the third step comprises the reorganization of the remaining β-sheet and of some unordered segments in the protein. The complete unfolding of the stable intermediate at higher temperatures also undergoes sequential events that ultimately end with the breaking of the H bonds between the two β-strands at the dimer interface.
Publication date
PublisherNational Academy of Sciences
AffiliationNRC Institute for Biodiagnostics; National Research Council Canada
Peer reviewedYes
NRC number1791
NPARC number9147680
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Record identifierafa2384b-7ba0-4eef-b3f3-363016df6780
Record created2009-06-25
Record modified2016-10-12
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