DINITROBENZENES STIMULATE ELECTRON FLUX WITHIN NEURONAL NITRIC OXIDE SYNTHASE IN THE ABSENCE OF CALMODULIN.
Efficient electron transfer and conversion ofL-arginine toL-citrulline and nitric oxide (NO•) by neuronal nitric oxide synthase (nNOS) requires calmodulin (CaM) binding. The present study focused on electron transfer ability of resting state CaM-free nNOS in presence of dinitrobenzene isomers (DNBs). NADPH oxidation (NADPHox) and acetylated cytochrome-creduction (AcCyt-cred) catalyzed by nNOS and the CaM binding sequence-deficient nNOS reductase construct (nNOS-FP) were estimates of total electron flux and [Formula: see text] production, respectively. All the DNBs (o-,m-,p-) independently stimulated rates of NADPHoxby CaM-free nNOS and by nNOS-FP in isomer- and concentration-dependent manner. Blocking nNOS heme by imidazole orL-arginine did not affect CaM-free nNOS-catalyzed NADPHoxstimulated by DNBs. This stimulated electron flux by DNBs did not support NO•formation by CaM-free nNOS. The DNBs, like FeCN, extract electrons from both FMN and FAD of the nNOS reductase domain. All three DNBs greatly stimulated nNOS and nNOS-FP catalyzed AcCyt-credthat was significantly inhibited by SOD demonstrating [Formula: see text] formation. Thus, in presence of DNBs, resting-state CaM-deficient nNOS efficiently transfers electrons generating [Formula: see text], inferring that additional metabolic roles for nNOS exist that are not yet explored.
Department of Biological Sciences, University of Texas at El Paso, 500 W University Ave, El Paso, TX 79968.
Chintamani N
CN
Joshi
Department of Physiology, Brody School of Medicine, East Carolina University, Greenville, NC 27834.
David A
DA
Tulis
Department of Biological Sciences, University of Texas at El Paso, 500 W University Ave, El Paso, TX 79968.
Richard T
RT
Miller
NADPH; nNOS; redox; superoxide