Substrate-specific Interactions with the Heme-bound Oxygen Molecule of Nitric-oxide Synthase

We report the characterization by resonance Raman spectroscopy of the oxygenated complex ((FeO2)-O-II) of nitric-oxide synthases of Staphylococcus aureus (saNOS) and Bacillus subtilis (bsNOS) saturated with N-omega-hydroxy-L-arginine. The frequencies of the nu(Fe-O) and nu(O-O) modes were 530 and 1135 cm(-1), respectively, in both the presence and absence of tetrahydrobiopterin. On the basis of a comparison of these frequencies with those of saNOS and bsNOS saturated with L-arginine (nu(Fe-O) at 517 cm(-1) and nu(O-O) at 1123 cm(-1)) and those of substrate-free saNOS (nu(Fe-O) at 517 and nu(O-O) at 1135 cm(-1)) (Chartier, F. J. M., Blais, S. P., and Couture, M. (2006) J. Biol. Chem. 281, 9953 9962), we propose two models that account for the frequency shift of nu(Fe-O) (but not nu(O-O)) upon N-omega-hydroxy-L-arginine binding as well as the frequency shift of nu(O-O) (but not nu(Fe-O)) upon L-arginine binding. The implications of these substrate-specific interactions with respect to catalysis by NOSs are discussed.