Evrard A, Zeghouf M, Fontecave M, Roby C, Covès J
Laboratoire de Résonance Magnétique en Biologie Métabolique, CEA-Grenoble and Université Joseph Fourier, Grenoble, France.
Eur J Biochem. 1999 Apr;261(2):430-7. doi: 10.1046/j.1432-1327.1999.00274.x.
SiR-FP60, the monomeric form of the Escherichia coli sulfite reductase flavoprotein component (SiR-FP), has been analysed by 31P-NMR spectroscopy. This protein was reported previously as a reliable simplified model for native SiR-FP [Zeghouf, M., Fontecave, M., Macherel, D., & Covès, J. (1998) Biochemistry 37, 6117-6123]. SiR-FP60 was examined in its native form, as a complex with NADP+ and after monoelectronic reduction either with NADPH or dithionite. In these latter cases, the stabilized FMN semiquinone radical offers a natural and internal paramagnetic probe. The paramagnetic effect of added manganese was also studied. In each case, the NMR parameters were extracted from digitalized data by a deconvolution procedure and compared with those obtained previously with cytochrome P450 reductase. Evolution of the NMR parameters and of calculated relaxation rate constants upon biochemical modifications of SiR-FP60 led us to propose that the reactive center is more compact than the one of cytochrome P450 reductase, with the redox components, FMN, FAD and NADPH, in a tighter spatial arrangement, close to the protein surface. This underlies some subtle differences between the two proteins for which a very similar overall structure is likely considering their common genetic origin and common operating cycle.
