Effect of the chaperone-like alpha-crystallin on the refolding of lysozyme and ribonuclease A.
Alpha-crystallin exhibits chaperone-like properties in preventing aggregation of proteins. We have studied the effect of alpha-crystallin on the refolding of denatured-disulfide intact and denatured-reduced lysozyme and RNase A. Alpha-crystallin does not have any effect on the refolding of both the denatured-disulfide intact enzymes. However, it inhibits the aggregation and oxidative renaturation of denatured-reduced lysozyme. Interestingly, it has no effect on the refolding of denatured-reduced RNase A. In order to probe the molecular basis of this differential behavior of alpha-crystallin towards lysozyme and RNase A, we have carried out circular dichroism and fluorescence studies on the refolding of denatured-reduced RNase A. It exhibits an extended conformation with little difference in the exposed hydrophobicity during the refolding process. We have earlier shown the presence of an aggregation-prone, refolding-competent, molten-globule-like intermediate on the refolding pathway of lysozyme. Alpha-crystallin binds to this intermediate, prevents its aggregation and inhibits its oxidative refolding. It was earlier believed that alpha-crystallin, unlike other chaperones, does not recognize intermediates on the refolding pathway but only recognizes intermediates on the unfolding pathway of proteins. Our present study clearly shows that it recognizes the refolding intermediates as well.
Raman
B
B
Centre for Cellular and Molecular Biology, Hyderabad, India.
Ramakrishna
T
T
Rao
C M
CM
D018833:Chaperonins / Q000378:metabolism*; D002942:Circular Dichroism; D003459:Crystallins / Q000378:metabolism*; D004220:Disulfides; D007700:Kinetics; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D011489:Protein Denaturation; D017510:Protein Folding; D012259:Ribonuclease, Pancreatic / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D003459:Crystallins; D004220:Disulfides; D012259:Ribonuclease, Pancreatic; D009113:Muramidase; D018833:Chaperonins