Detection of a Nitric Oxide Synthase Possibly Involved in the Regulation of theRhodococcussp R312 Nitrile Hydratase

Crude homogenates from Rhodococcus sp 312 catalyze the conversion of L-arginine into L-citrulline and NO2-, the usual oxidation product of NO under aerobic conditions. They also catalyze the conversion of N-omega-hydroxy-L-arginine (NOHA) into L-citrulline and NO2- with similar rates (10-15 and 100-150 nmol of product min(-1) . (mg of protein)(-1) respectively for the crude homogenate and for a fraction obtained from ammonium sulfate precipitation). L-citrulline formation is strongly inhibited by classical inhibitors of mammalian nitric oxide synthases (NOSs) such as N-omega-methyl-L-arginine (NMA) and thio-L-citrulline (TC). Finally, the lack of inhibitory effects of EGTA, a classical inhibitor of constitutive mammalian NOSs, and the specific immunodetection of a 100 kD protein from Rhodococcus cytosol by an antibody raised against human inducible NOS, is in favor of the presence of a NOS similar to inducible mammalian NOSs in Rhodococcus sp 312. This NOS should be responsible for the NO-dependent inactivation of Rhodococcus Nitrile Hydratase (NHase) in the absence of light; it could regulate the activity of the latter enzyme. (C) 1998 Academic Press.