Document Type
Article
Publication Date
2013
Publication Title
Biochemical Journal
Abstract
The NOS (nitric oxide synthase; EC 1.14.13.39) enzymes contain a C-terminal flavoprotein domain [NOSred (reductase domain of NOS)] that binds FAD and FMN, and an N-terminal oxygenase domain that binds haem. Evidence suggests that the FMN-binding domain undergoes large conformational motions to shuttle electrons between the NADPH/FAD-binding domain [FNR (ferredoxin NADP-reductase)] and the oxygenase domain. Previously we have shown that three residues on the FMN domain (Glu(762), Glu(816) and Glu(819)) that make charge-pairing interactions with the FNR help to slow electron flux through nNOSred (neuronal NOSred). In the present study, we show that charge neutralization or reversal at each of these residues alters the setpoint [K-eq(A)], of the NOSred conformational equilibrium to favour the open (FMN-deshielded) conformational state. Moreover, computer simulations of the kinetic traces of cytochrome c reduction by the mutants suggest that they have higher conformational transition rates (1.5-4-fold) and rates of interflavin electron transfer (1.5-2-fold) relative to wild-type nNOSred. We conclude that the three charge-pairing residues on the FMN domain govern electron flux through nNOSred by stabilizing its closed (FMN-shielded) conformational state and by retarding the rate of conformational switching between its open and closed conformations.
Recommended Citation
Haque, Mohammad Mahfuzul; Bayachou, Mekki; Fadlalla, Mohammed A.; Durra, Deborah; and Stuehr, Dennis J., "Charge-Pairing Interactions Control The Conformational Setpoint and Motions of The FMN Domain in Neuronal Nitric Oxide Synthase" (2013). Chemistry Faculty Publications. 304.
https://engagedscholarship.csuohio.edu/scichem_facpub/304
DOI
10.1042/BJ20121488
Version
Postprint
Volume
450
Comments
This work was supported by the National Institutes of Health [grant numbers GM51491 and HL58883 (to D.J.S.)]