Antibody as an Immunological Probe for Studying the Refolding of Bovine Serum Albumin I. THE CATALYSIS OF REOXIDATION OF REDUCED BOVINE SERUM ALBUMIN BY GLUTATHIONE AND A DISULFIDE INTERCHANGE

We have used an immunochemical approach to study the refolding of bovine serum albumin. Using antibody as a probe for return of native structure, we have been able to demonstrate the regeneration of native structure at several sites on the surface of the molecule. Using this technique, we have shown that the rate of refolding of reduced bovine serum albumin catalyzed by either glutathione or rat liver disulfide interchange enzyme is greater than the rate of air reoxidation of albumin. The half-regeneration times for albumin, however, are substantially greater than those obtained with smaller proteins that have fewer disulfide bonds. We have also demonstrated that the reoxidized monomers isolated at the end of the refolding process are immunologically identical to native monomers. In addition, the tryptophan fluorescence emission maxima were the same as that of the native monomers.