CATALYTIC MECHANISM OF DSBA AND ITS COMPARISON WITH THAT OF PROTEIN DISULFIDE-ISOMERASE

The mechanism of action of the bacterial periplasm protein DsbA in int roducing disulfide bonds into proteins was studied by its action on a model disordered peptide containing only two cysteine residues. Most o f the reactions between the various thiol and disulfide forms of the p eptide and of DsbA could be measured directly. All those involving Dsb A occurred 10(2)-10(6) times more rapidly than is normally observed be tween other typical thiols and disulfides; DsbA apparently stabilizes the transition state of thiol-disulfide exchange. The reactions betwee n DsbA and the peptide were even more rapid, and they were constrained to occur at only one sulfur atom of disulfide bonds involving the pep tide. Both observations indicate that noncovalent binding interactions also occur between DsbA and the peptide, and the expected effect of b inding between reactants on rates of reaction was quantified. Small qu antities of DsbA had catalytic effects on the reaction between the pep tide and glutathione, similar to those observed previously with the eu karyotic catalyst protein disulfide isomerase. The known reactions of DsbA could account quantitatively for these effects and indicated that the apparent catalysis was the result of the separate and sequential rapid reactions of the peptide and of glutathione at the active site o f DsbA. DsbA did not catalyze the conformational changes involved in f orming an intramolecular disulfide bond in the peptide; its catalytic effects were simply due to its rapid participation in thiol-disulfide exchange reactions. Protein disulfide isomerase is likely to function very similarly to DsbA.