Endothelial Nitric-oxide Synthase

• DOI: 10.1074/jbc.271.24.14631
• Publication Date: 2002-07-26T14:52:05Z
• Authors: Pei-Feng Chen; Ah-Lim Tsai; Vladimir Berka; Kenneth K. Wu
• Keywords: Molecular mass; Cytochrome P450 reductase

A baculovirus system was used to express the oxygenase and reductase domains of human endothelial nitric-oxide synthase (ecNOS) as distinct proteins. The domain (residues 1–491) expressed using a vector containing His6 tag at N terminus. purified had an apparent molecular mass ∼54 kDa, retained ability bind L-arginine form ferrous CO complex. 492-1244) ∼82 kDa catalyze NADPH-dependent cytochrome c reduction, which enhanced 10-fold by presence Ca2+/calmodulin. Both exhibited immunoreactivity rabbit anti-ecNOS IgG. NOS activity successfully reconstituted mixing two domains. These results demonstrate for first time that ecNOS are catalytically intact can be in vitro.