Use of the phase diagram method to analyze the protein unfolding-refolding reactions: fishing out the "invisible" intermediates

Journal of Proteome Research
Irina M KuznetsovaVladimir N Uversky

Abstract

Partially folded conformations are important players in protein self-organization, function, and misfolding, thus attracting the intensive and constant attention of researchers. Different conformational states of proteins can be easily discriminated from one another using the multiparametric approach, which usually involves the application of several physicochemical methods sensitive to the various structural levels of a protein molecule. This approach gives the most complete information about the structural changes in protein during its de- and renaturation. However, in many cases researches are interested in a brief and simple test for the appearance of an intermediate state. The binding of the hydrophobic fluorescent probe ANS represents one of the well-established techniques, allowing the retrieval of such information. We are introducing another approach, based on the analysis of the spectroscopic data in a form of spectral diagrams. The essence of this method is to build the Ilambda1, versus Ilambda2 dependence, where Ilambda1 and Ilambda2 are the spectral intensity values (e.g., fluorescence, CD, FTIR, absorbance, etc.) measured on wavelengths lambda1 and lambda2. We are showing that this approach gives the detailed descr...Continue Reading

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Citations

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