PROBING NUCLEOTIDE DEPENDENT CONFORMATIONAL CHANGE IN THE NITROGENASE FE PROTEIN

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. During nitrogenase catalysis, the iron protein and molybdenum-iron protein associate and dissociate in a manner resulting in the hydrolysis of two molecules of MgATP and the transfer of at least one electron to the MoFe protein. These nucleotide dependent conformational changes are analogous to a number of other examples of biochemical signal transduction machinery and therefore probing the role of nucleoside triphosphate binding and hydrolysis in nitrogenase is of broad significance. There is considerable biochemical and spectroscopic evidence including small angle x-ray scattering studies that have implicate that the Fe protein undergoes conformational changes during nucleotide displacement (MgATP for MgADP or nucleoside triphosphate hydrolysis. We have recently determined the structure of a site-direct variant of the Fe protein that has a number of biochemical and spectroscopic features in the absence of nucleotide that resemble the native Fe protein in the presence of bound MgATP. The most significant structural differences that separate the Fe protein variant and the aforementioned structures of the Fe protein are largely manifested in a rigid body reorientation of the Fe protein subunits with respect to one another. Each of the Fe protein subunits is reoriented by a rigid-body rotation with respect to one another on the order of sixty to eighty degrees relative to the previously characterized Fe protein structures. Our calculation using the crystallographic coordinates indicates that the site-direct variant has a radius of gyration approximately 2.2 ¿ larger than that of the native Fe protein despite the missing residues in the variant. This result apparently contradicts the previous SAXS results on the native protein upon MgATP binding [Chen et al., (1994) J. Biol. Chem. 269, 3290]. We would like to revisit SAXS of the native Fe protein in nucleotide bound states to examine the validity of our hypothetical model for the conformational changes the Fe protein undergoes.