Effect of solution viscosity on intraprotein electron transfer between the FMN and heme domains in inducible nitric oxide synthase

Abstract

The FMN-heme intraprotein electron transfer (IET) kinetics in a human inducible NOS (iNOS) oxygenase/FMN construct were determined by laser flash photolysis as a function of solution viscosity (1.0-3.0 cP). In the presence of ethylene glycol or sucrose, an appreciable decrease in the IET rate constant value was observed with an increase in the solution viscosity. The IET rate constant is inversely proportional to the viscosity for both viscosogens. This demonstrates that viscosity, and not other properties of the added viscosogens, causes the dependence of IET rates on the solvent concentration. The IET kinetics results indicate that the FMN-heme IET in iNOS is gated by a large conformational change of the FMN domain. The kinetics and NOS flavin fluorescence results together indicate that the docked FMN/heme state is populated transiently.