Interferon-gamma-inducible murine macrophage nitric oxide synthase: studies on the mechanism of inhibition by imidazole agents.

Citrulline formation by the interferon-gamma/lipopolysaccharide-inducible murine macrophage nitric oxide synthase is inhibited reversibly by imidazole, 1-phenylimidazole, 4-phenylimidazole, and 2-phenylimidazole with IC50 values of 40 microM, 6 microM, 225 microM, and > 1 mM, respectively. 1-Phenylimidazole inhibited the maximal velocity of citrulline formation but did not alter the concentration of arginine providing half-maximal activity. 1-Phenylimidazole inhibited citrulline formation by the murine macrophage nitric oxide synthase competitively versus (6R)-5,6,7,8-tetrahydro-L-biopterin (THB) with a Ki value of 0.7 microM, but inhibited citrulline formation by Ca(2+)-calmodulin-dependent nitric oxide synthase from GH3 pituitary cells noncompetitively versus THB with a Ki value of 40 microM. Imidazole inhibited citrulline formation by the murine macrophage nitric oxide synthase noncompetitively versus THB with a Ki value of 48 microM. Neither imidazole nor 1-phenylimidazole inhibited the cytochrome c reductase activity of murine macrophage nitric oxide synthase at concentrations 100-fold higher than their IC50 values for inhibiting citrulline formation. The antifungal imidazoles miconazole, ketoconazole, and clotrimazole did not inhibit either citrulline formation or cytochrome c reduction by murine macrophage nitric oxide synthase at concentration as high as 200 microM. Ca(2+)-calmodulin-dependent nitric oxide synthase from GH3 pituitary cells exhibited a Kact for THB of 80 nM, while the inducible murine macrophage nitric oxide synthase exhibited a Kact of 8 microM.