Probing the MgATP-bound conformation of the nitrogenase Fe protein by solution small-angle X-ray scattering.
The MgATP-bound conformation of the Fe protein of nitrogenase from Azotobacter vinelandii has been examined in solution by small-angle X-ray scattering (SAXS) and compared to existing crystallographically characterized Fe protein conformations. The results of the analysis of the crystal structure of an Fe protein variant with a Switch II single-amino acid deletion recently suggested that the MgATP-bound state of the Fe protein may exist in a conformation that involves a large-scale reorientation of the dimer subunits, resulting in an overall elongated structure relative to the more compact structure of the MgADP-bound state. It was hypothesized that the Fe protein variant may be a conformational mimic of the MgATP-bound state of the native Fe protein largely on the basis of the observation that the spectroscopic properties of the [4Fe-4S] cluster of the variant mimicked in part the spectroscopic signatures of the native nitrogenase Fe protein in the MgATP-bound state. In this work, SAXS studies reveal that the large-scale conformational differences between the native Fe protein and the variant observed by X-ray crystallography are also observed in solution. In addition, comparison of the SAXS curves of the Fe protein nucleotide-bound states to the nucleotide-free states indicates that the conformation of the MgATP-bound state in solution does not resemble the structure of the variant as initially proposed, but rather, at the resolution of this experiment, it resembles the structure of the nucleotide-free state. These results provide insights into the Fe protein conformations that define the role of MgATP in nitrogenase catalysis.
Sarma
Ranjana
R
Department of Chemistry and Biochemistry, Montana State University, Bozeman, Montana 59717, USA.
Mulder
David W
DW
Brecht
Eric
E
Szilagyi
Robert K
RK
Seefeldt
Lance C
LC
Tsuruta
Hiro
H
Peters
John W
JW
D000255:Adenosine Triphosphate / Q000378:metabolism*; D016948:Azotobacter vinelandii / Q000201:enzymology; D010088:Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction
D000255:Adenosine Triphosphate; D010088:Oxidoreductases; C029130:nitrogenase reductase