Important role of tetrahydrobiopterin in NO complex formation and interdomain electron transfer in neuronal nitric-oxide synthase

Neuronal nitric-oside synthase (nNOS) is composed of a heme oxygenase domai n and a flavin-bound reductase domain. Ca2+/calmodulin (CaM) is essential f or interdomain electron transfer during catalysis, whereas the role of the catalytically important cofactor, tetrahydrobiopterin (H4B) remains elusive , The product NO appears to bind to the heme and works as a feedback: inhib itor, The present study shows that the Fe3+-NO complex is reduced to the Fe 2+-NO complex by NADPH in the presence of both L-Arg and H4B even in the ab sence of Ca2+/CaM. The complex could not be fully reduced in the absence of H4B under any circumstances. However, dihydrobioptepin and N-G-hydroxy-L-A RG, respectively could be substituted for H4B and L-Arg, respectively, No d irect correlation could be found between redox potentials of the nNOS heme and the observed reduction of the Fe3+-NO complex, Thus, sur data indicate the importance of the pterin binding to the active site structure during th e reduction of the NO-heme complex by NADPH during catalytic turnover. (C) 2001 Academic Press.