Unfolding/Refolding Studies on Proteins
Introduction:
Considerable attention is currently being paid to the unfolding/refolding mechanism of protein molecules in respect of three-dimensional structure formation of polymer molecules, as well as the maturation of a protein newly synthesized in cells and the production of an active form of recombinant protein by using the DNA technology. Even if a recombinant protein maintains its biological activities, conformational change in recombinant protein as compared with native protein lead to anaphylaxis. From several in vitro studies in which renaturation of denatured protein was attempted, it has been assumed that, in general, a relatively small protein with a molecular weight below 20,000 could reversibly refold from the denatured state. Ribonuclease A,alpha-lactalbumin and bovine pancreatic trypsin inhibitor are examples of those proteins that were successfully refolded. The unfolding/refolding studies on these model proteins have been chiefly focused on the folding pathway, the mechanism for stabilizing the folding intermediates, and the rate-limiting step in folding.
Purpose of our study is as follows:
1) Evaluate whether a relatively small protein after renaturation from the denatured form could regain the native structure in all local moieties within the molecule.
2) Establish the method to achieve the complete refolding of a relatively small protein from denatured state.
3) Elucidate the disulfide folding pathway of a relatively small protein.
In this research projject, we are collaborating with Drs. Shuichi Kaminogawa and Akio Ametani of the University of Tokyo.
Reference
Hattori, M.; Hiramatsu, K.; Kurata, T.; Nishiura, M.; Takahashi, K.; Ametani, A.; Kaminogawa, S., Complete Refolding of Bovine beta-Lactoglobulin Requires Disulfide Bond Formation under Strict Conditions. Biochim. Biophys. Acta 2005, 1752, 154-165. [Abstract]
Hattori, M.; Ametani, A.; Katakura, Y.; Shimizu, M.; Kaminogawa, S. Unfolding/Refolding Studies on Bovine beta-Lactoglobulin with Monoclonal Antibodies as Probes: Does a Renatured Protein Completely Refold ? J. Biol. Chem. 1993, 268, 22414-22419.
[Abstract]
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