EVIDENCE FOR THE PRESENCE OF AN UNUSUAL NITRIC-OXIDE-PRODUCING AND CITRULLINE-PRODUCING ENZYME IN RAT-KIDNEY

We have found an enzymatic activity obtained from rat kidney capable o f producing citrulline and NOx (nitrate and nitrite) which was resista nt to inhibition by conventional arginine analogues. This enzyme activ ity does not require any calcium or calmodulin and was found to be ind uced during pregnancy. This unique enzyme was found to be tissue and s pecies specific. Another unique feature of this enzyme is that it did not bind to 2'5'-ADP-sepharose under standard conditions. Western blot analysis of the 100,000 g kidney supernatant using monoclonal antibod y for macrophage inducible nitric oxide synthase failed to produce a b and for inducible nitric oxide synthase. HPLC and capillary ion analys is for nitrate and nitrite (NOx) showed clear peaks for [H-3] L-citrul line and NOx, respectively, which were not changed either in the absen ce of calcium and calmodulin or in the presence of 300 mu M S-ethyliso thiourea, which has been shown to be a very potent and selective inhib itor of inducible nitric oxide synthase with a Ki of about 14.7 nM. Th ese results suggest the possible existence of another isoform of nitri c oxide synthase with very distinct properties from the known isoforms . (C) 1997 Academic Press.