Using dysprosium complexes to probe the nitrogenase paramagnetic centers.
EPR progressive power saturation techniques were used to monitor relaxation enhancement of the nitrogenase paramagnetic centers produced by interaction with Dy3+ complexes. Three models are presented for the relationship between the degree of enhancement and the distance of closest approach of Dy3+ to the intrinsic metal cluster. In the first model, the perturbing dysprosium ions are represented as a single average site. The second and third models are variations of the treatment of Innes and Brudvig [(1989) Biochemistry 28, 1116-1125] and assume the unknown protein to be spherical with Dy3+ dispersed either randomly over the surface of the protein or randomly in solution. Using these models, the distance of closest approach of the Dy3+ complex to the [4Fe-4S] cluster in the Fe-protein from Azotobacter vinelandii was determined to be 5.0-6.5 A. Similarly, the distance of closest approach to FeMoco in the MoFe-protein was determined to be 0.0-1.2 A, which, when corrected to the fact that FeMoco exists as an S = 3/2 spin state, indicates that the distance is > or = 6 A. These distances did not change when (1) either protein was in the presence of the other, (2) both proteins were cross-linked to each other, or (3) the Fe-protein from A. vinelandii was mixed with the MoFe-protein from Clostridium pasteurianum. On the other hand, formation of the inactive complex of the Fe-protein from C. pasteurianum with the MoFe-protein from A. vinelandii blocked dysprosium-induced relaxation enhancement, implying that each component protein overlaps the metal cluster in the complementing protein.
Department of Chemistry, Louisiana State University, Baton Rouge 70803-1804.
M E
ME
Oliver
B J
BJ
Hales
D016948:Azotobacter vinelandii / Q000201:enzymology; D004419:Dysprosium; D004578:Electron Spin Resonance Spectroscopy; D008956:Models, Chemical; D008983:Molybdoferredoxin / Q000737:chemistry; D009591:Nitrogenase / Q000737:chemistry*; D010088:Oxidoreductases
D008983:Molybdoferredoxin; D004419:Dysprosium; D010088:Oxidoreductases; D009591:Nitrogenase; C029130:nitrogenase reductase