Mitochondrial NOS activity: Regulation by substrates and mitochondrial metabolic states

BOVERIS A; ZAOBORNYJ T; ALVAREZ S; VALDEZ LB

Santa Bárbara, California, USA

Congreso; Annual Meeting of Oxygen Club of California. 2004 World Congress: Oxidants & Antioxidants in Biology; 2004

Oxygen Club of California

Nitric oxide (NO) has been recognized as a signaling molecule that acts both as an intercellular messenger and as an intracellular regulator. This molecule is produced in mitochondria by the enzymatic co-oxidation of L-arginine and NADPH by O2 to yield NADP, L-citrulline and NO, in a reaction catalyzed by a mitochondrial nitric oxide synthase (mtNOS). The aim of this work was to study the regulatory effects of L-arginine, O2, pH and metabolic states on mtNOS activity from different rat organs: heart, liver, kidney and diaphragm. The optimum pH for mtNOS activity was in the 6.5-7.4 range for submitochondrial membranes (SMM) of the organs tested, with a NO production of about 0.7-1.4 nmol/min.mg protein. Mitochondria seem to contribute with about 67% (heart), 37% (liver), 30% (kidney) and 24% (diaphragm) of the total cellular NO production. The study of NO production by SMM as a function of L-arginine concentration showed a hyperbolic response. The apparent L-arginine KM were 36 mM (heart), 70 mM (liver), 4 mM (kidney) and 37 mM (diaphragm). Values for the apparent KM for O2 were 3.3 mM (heart), 40 mM (liver), 37 mM (kidney) and 4.6 mM (diaphragm). The state 4/state 3 transition regulates mtNOS activity and NO release in coupled respiring mitochondria. As a whole, NO production rates at state 3 were 40-50% lower than at state 4. The study of the substrate and state 4/state 3 transition dependence of mtNOS activity, provides the basis for the understanding of the physiological relevance of NO production in mitochondria.