title: Intraprotein electron transfer between the FMN and heme domains in endothelial nitric oxide synthase holoenzyme.
abstract: Intraprotein electron transfer (IET) from flavin mononucleotide (FMN) to heme is an essential step in nitric oxide (NO) synthesis by NO synthase (NOS). The IET kinetics in neuronal and inducible NOS (nNOS and iNOS) holoenzymes have been previously determined in our laboratories by laser flash photolysis [reviewed in: C.J. Feng, G. Tollin, Dalton Trans., (2009) 6692-6700]. Here we report the kinetics of the IET in a bovine endothelial NOS (eNOS) holoenzyme in the presence and absence of added calmodulin (CaM). The IET rate constant in the presence of CaM is estimated to be ~4.3s(-1). No IET was observed in the absence of CaM, indicating that CaM is the primary factor in controlling the FMN-heme IET in the eNOS enzyme. The IET rate constant value for the eNOS holoenzyme is approximately 10 times smaller than those obtained for the iNOS and CaM-bound nNOS holoenzymes. Possible mechanisms underlying the difference in IET kinetics among the NOS isoforms are discussed. Because the rate-limiting step in the IET process in these enzymes is the conformational change from input state to output state, a slower conformational change (than in the other isoforms) is most likely to cause the slower IET in eNOS.
authors.affiliation: College of Pharmacy, University of New Mexico, Albuquerque, NM 87131, USA. cfeng@salud.unm.edu
authors.forename: Changjian
authors.initials: C
authors.lastname: Feng
authors.forename: Valentina
authors.initials: V
authors.lastname: Taiakina
authors.forename: Dipak K
authors.initials: DK
authors.lastname: Ghosh
authors.forename: J Guy
authors.initials: JG
authors.lastname: Guillemette
authors.forename: Gordon
authors.initials: G
authors.lastname: Tollin
mesh_terms: D000818:Animals; D002417:Cattle; D004579:Electron Transport / Q000502:physiology*; D005486:Flavin Mononucleotide / Q000737:chemistry* / Q000378:metabolism; D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D020035:Holoenzymes / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D052250:Nitric Oxide Synthase Type III / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects; D055668:Photochemical Processes; D010782:Photolysis; D017434:Protein Structure, Tertiary / Q000502:physiology; D056928:X-Ray Absorption Spectroscopy
chemical_list: D020035:Holoenzymes; D007527:Isoenzymes; D006418:Heme; D005486:Flavin Mononucleotide; D052250:Nitric Oxide Synthase Type III