title: Cross-linking of nitrogenase components. Structure and activity of the covalent complex.
abstract: The nitrogenase complex from Azotobacter vinelandii is composed of the MoFe protein (Av1), an alpha 2 beta 2 tetramer, and the Fe protein (Av2), a gamma 2 dimer. During turnover of the enzyme, electrons are transferred from Av2 to Av1 in parallel with the hydrolysis of MgATP. Using the cross-linking reagent, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, we have identified some of the properties of the complex between the two components. The cross-linking reaction was highly specific yielding a single apparent Mr = 97,000 protein. The amount of cross-linked product was essentially independent of whether MgATP or MgADP were in the reaction. Also, the amount was maximum at high ratios of Av2 to Av1. The Mr = 97,000 protein was characterized by amino acid analysis and Edman degradation and was found to be consistent with a 1:1 complex of an Av2 gamma subunit and an Av1 beta subunit (the amino terminal serine subunit). The complex was no longer active in the nitrogenase reaction which supports, but does not prove, the requirement for dissociation of the complex after each electron transferred. Nitrogenase activity and cross-linking were inhibited in an identical way by NaCl, which suggests that electrostatic forces are critical to the formation of the electron transfer complex.
authors.affiliation: Department of Biochemistry, University of Minnesota, Minneapolis 55455.
authors.forename: A H
authors.initials: AH
authors.lastname: Willing
authors.forename: M M
authors.initials: MM
authors.lastname: Georgiadis
authors.forename: D C
authors.initials: DC
authors.lastname: Rees
authors.forename: J B
authors.initials: JB
authors.lastname: Howard
mesh_terms: D000596:Amino Acids / Q000032:analysis; D001395:Azotobacter / Q000201:enzymology*; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009591:Nitrogenase / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D046911:Macromolecular Substances; D009591:Nitrogenase; D005022:Ethyldimethylaminopropyl Carbodiimide