Both the isomerase and chaperone activities of protein disulfide isomerase are required for the reactivation of reduced and denatured acidic phospholipase A(2)

The spontaneous reactivation yield of acidic phospholipase A(2) (APLA(2)), a protein containing seven disulfide bonds, after reduction and denaturation in guanidine hydrochloride is very low, Protein disulfide isomerase (PDI) markedly increases the reactivation yield and prevents the aggregation of APLA(2) during refolding in a redox buffer containing GSH and GSSG. S-methylated PDI (mPDI), with no isomerase but as nearly full chaperone activity as native PDI, has no effect on either the reactivation or aggregation of APLA(2). However, the simultaneous presence of PDI and mPDI in molar ratios to APLA(2) of 0.1 and 0.9 respectively fully reactivates the denatured enzyme, as does PDI alone at a ratio of 1. At ratios of 0.1 and 0.15 respectively, they completely suppress APLA(2) aggregation, as does PDI alone at a ratio of 0.25, Moreover, delayed addition of PDI to the refolding buffer greatly diminished the reactivation yield of APLA(2), but this deteriorating effect can be alleviated markedly by the presence of mPDI in the refolding buffer, Without GSSG, mPDI prevents the aggregation of APLA(2) during refolding, It is proposed that the in vitro action of PDI as a foldase consists of both isomerase and chaperone activities, and the latter activity can be fully replaced by mPDI.