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Control Of Electron Transfer And Catalysis In Neuronal NOS By A Hinge Connecting The FMN And FNR Domains
Author(s) -
Haque Mohammad M.,
Fadlalla Mohammed,
Aulak Kulwant,
Ghosh Arnab,
Stuehr Dennis
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.573.7
Subject(s) - hinge , electron transfer , chemistry , enzyme , flavin mononucleotide , flavin group , electron transport chain , flavoprotein , biophysics , stereochemistry , crystallography , biochemistry , photochemistry , biology , physics , classical mechanics
In nitric oxide synthase enzymes (NOSs) two flexible hinge elements connect the FMN domain to the rest of the enzyme and may guide FMN domain interactions with its partner FNR and NOSoxy domains to enable the electron transfers required during catalysis. In our previous study, we found that incorporating the nNOS FMN‐FNR hinge into eNOS increased its NO synthesis activity 4‐fold, decreased uncoupled NADPH oxidation, and supported faster heme reduction (kr), implying that the composition and/or length of the FMN‐FNR hinge is important for determining electron transfer and catalysis. To more systematically investigate the role of the FMN‐FNR hinge we constructed mutants in nNOS that either shorten or lengthen the hinge by 2, 4, and 6 residues. We found that FMN‐FNR hinge length helps determine electron flux through the nNOS flavoprotein domain. Shortening the hinge inhibited electron flux through both the calmodulin (CaM)‐free and CaM‐bound enzymes, while lengthening the hinge relieved the repression on electron flux that is normally present in the CaM‐free enzyme, and has either no impact or was deleterious for electron flux through the CaM‐bound enzyme. The nNOS kr was also sensitive to FMN‐FNR hinge length. Small changes in hinge length were tolerated, while changes beyond 2 or 4 residues were deleterious to kr, and also decoupled nNOS NADPH consumption from NO synthesis. Supported by NIH GM51941 & HL63744.