IBIMOL   23987
INSTITUTO DE BIOQUIMICA Y MEDICINA MOLECULAR PROFESOR ALBERTO BOVERIS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Mitochondrial nitric oxide production supported by reverse electron transfer
Autor/es:
ZAOBORNYJ T; BOMBICINO SS; BOVERIS A; BOMBICINO SS; BOVERIS A; IGLESIAS DE; IGLESIAS DE; VALDEZ LB; VALDEZ LB; ZAOBORNYJ T
Lugar:
Tucumán
Reunión:
Congreso; III Latin American Federation of Biophysical Societies (LAFeBS), IX IberoAmerican Congress of Biophysics and XLV Reunion Annual SAB; 2016
Institución organizadora:
Sociedad Argentina de Biofísica
Resumen:
The aim of this work was to study mitochondrial NO production using phosphorylating electron transfer particles (ETPH) obtained from bovine heart. In these vesicles, the mitochondrial inner membrane has a reverse orientation with the NADH dehydrogenase center of complex I and the F1-ATPase exposed to the solutes in the surrounding medium. In our experimental conditions, 60% of the particles are in reverse status, estimated both through the O2 consumption rate using NADH as substrate and the NADH-O2 oxidoreductase activity, in the presence or absence of cytochrome c3+. ETPH produced 1.2 ± 0.1 nmol NO. min-1. mg protein-1 through the reaction catalyzed by mitochondrial nitric oxide synthase (mtNOS). These particles showed a succinate-NAD+ reductase activity of 64 ± 3 nmol NADH. min-1. mg protein-1 sustained by reverse electron transfer (RET) at expenses of ATP and succinate. The same particles, without NADPH and in conditions of RET produced 0.97 ± 0.07 nmol NO. min-1. mg protein-1. Rotenone inhibited NO production supported by RET measured in ETPH and in coupled mitochondria, but did not decrease the activity of recombinant nNOS, indicating that the inhibitory effect of rotenone on NO production is due to an electron flow inhibition and not to a direct action on mtNOS structure. NO production sustained by RET corresponds to 20% of the total amount of NO released from heart coupled mitochondria. A mitochondrial fraction enriched in complex I produced 1.7 ± 0.2 nmol NO. min-1. mg protein-1 and reacted with anti 75 kDa complex I subunit and anti-nNOS antibodies, suggesting that complex I and mtNOS are located contiguously. These data show that mitochondrial NO production can be supported by RET, and suggest that mtNOS is next to complex I, reaffirming the idea of a functional association between these proteins.This work was supported by UBA (UBACYT 20020090200393, 20020110100140, 20020100100606), ANPCyT (PICT 2008-1138; 2010-0844; PICT 2014-0964), and CONICET (PIP 11220080100688, PIP 11220110100444).