Aminoguanidine-mediated inactivation and alteration of neuronal nitric-oxide synthase

It is established that aminoguanidine (AG) is a metabolism-based inactivato r of the three major isoforms of nitric-oxide synthase. AG is thought to be of potential use in diseases, such as diabetes, where pathological overpro duction of NO is implicated. We show here that during the inactivation of n euronal nitric-oxide synthase (nNOS) by AG that the prosthetic heme is alte red, in part, to dissociable and protein-bound adducts, The protein-bound h eme adduct is the result of cross-linking of the heme to residues in the ox ygenase domain of nNOS. The dissociable heme product is unstable and revert s back to heme upon isolation. The alteration of the heme is concomitant wi th the loss in the ability to form the ferrous-CO complex of nNOS and accou nts for at least two-thirds of the activity loss. Studies with [C-14]AG ind icate that alteration of the protein, in part on the reductase domain of nN OS, also occurs but at low levels. Thus, heme alteration appears to be the major cause of nNOS inactivation. The elucidation of the mechanism of inact ivation of nNOS will likely lead to a better understanding of the in vivo e ffects of NOS inhibitors such as AG.