Selective recognition of glutathiolated aldehydes by aldose reductase

In this study, the selectivity and specificity of aldose reductase (AR) for glutathionyl aldehydes was examined, Relative to free aldehydes, AR was a more efficient catalyst for the reduction of glutathiolated aldehydes, Redu ction of glutathionyl propanal [gamma Glu-Cys(propanal)-Gly] was more effic ient than that of Gly-Cys(propanal)-Gly and gamma-aminobutyric acid-Cys(pro panal)-Gly suggesting a possible interaction between alpha-carboxyl of the conjugate and AR. Two active site residues, Trp20 or Ser302, were identifie d by molecular modeling as potential sites of this interaction, Mutations c ontaining tryptophan-to-phenylalanine (W20F) and serine-to-alanine (S302A) substitutions did not significantly affect reduction of free aldehydes but decreased the catalytic efficiency of AR for glutathiolated aldehydes, Comb ined mutations indicate that both Trp20 and Ser302 are required for efficie nt catalysis of the conjugates. The decrease in efficiency due to W20F muta tion with glutathionyl propanal was not observed with gamma-aminobutyric-Cy s(propanal)-Gly or Gly-Cys-(propanal)-Gly, indicating that Trp20 is involve d in binding the cc-carboxyl of the conjugate, The effect of the S302A muta tion was less severe when gamma Glu-Cys(propanal)-Glu rather than glutathio nyl propanal was used as the substrate, consistent with an interaction betw een Ser302 and Gly-3 of the conjugate. These observations suggest that glut athiolation facilitates aldehyde reduction by AR and enhances the range of aldehydes available to the enzyme. Because the N-terninal carboxylate is un ique to glutathione, binding of the conjugate with the alpha-carboxyl facin g the bottom of the alpha/beta-barrel may assist in the exclusion of unrela ted peptides and proteins.