Functional implication of disulfide bond, Cys250 -Cys283, in bovine chymosin

The reduction, carboxymethylation and mercuration of disulfide bond, Cys250-Cys283, located on the surface of bovine chymosin molecule resulted in the loss of about 25% of enzyme activity, suggesting that Cys250-Cys283 is not intimately involved in catalytic mechanism. Cys250 and Cys283 were substituted with Asp. and Ser. by site- directed mutagenesis of the structural gene coding for bovine prochymosin B. All three mutants (C250D/C283S, C250D, C283S) failed to be activated to chymosin in acid, indicating that Cys250-Cys283 might have some contribution to the correct refolding of the unfolded prochymosin.