PROTEIN DISULFIDE ISOMERASE-CATALYZED RENATURATION OF RIBONUCLEASE-A MODIFIED BY S-THIOLATION WITH GLUTATHIONE AND CYSTEINE

Renaturation of modified ribonuclease A by protein disulfide isomerase was studied. The renaturation rate of fully S-thiolated ribonuclease A with glutathione, namely, ribonuclease A-glutathione mixed disulfide (RNase-SG) containing 8 moles of glutathione per mole of ribonuclease A (RNase-SG8), by protein disulfide isomerase (PDI) was more than thr ee times faster than those of fully S-thiolated RNase with L-cysteine and scrambled ribonuclease A. Renaturation of RNase-SG species contain ing 7 or less glutathione was slower than that of RNase-SG8. These dat a seems to favor the hypothesis that S-thiolation of nascent proteins with glutathione may occur in the folding process during protein synth esis. The applicability of the present method consisted of chemical S- thiolation and PDI-catalyzed renaturation to the in vitro folding of r ecombinant cysteine-containing proteins is discussed.