Nitrogenase is a complex enzyme: It contains an iron (Fe) protein dimer and a molybdenum-iron (MoFe) protein α2B2 tetramer, and it catalyzes reduction of dinitrogen to ammonia. The Fe protein transfers electrons to the MoFe protein and also is involved in the biosynthesis of a Mo-7Fe-9S-homocitrate (FeMoco) cluster and in the insertion of this cofactor into the α subunits of the MoFe protein.
Ribbe and Burgess show that insertion of this cofactor, which forms the binding site for dinitrogen, requires the chaperone GroEL. They used a mutant strain of Azobacter vinelandii that is defective in FeMoco insertion to generate a FeMoco-deficient MoFe protein. Addition of a crude extract together with MgATP and wild-type Fe protein restored full activity in vitro to partially purified FeMoco-deficient MoFe protein. The component required for FeMoco insertion was purified and identified as GroEL by NH2-terminal sequencing and antibody cross-reactivity. Yet GroEL, MgATP, and Fe protein were not sufficient for FeMoco insertion into purified FeMoco-deficient MoFe protein, suggesting that other factors also help to escort this metal cluster. — VV
Proc. Natl. Acad. Sci. U.S.A.98, 5521 (2001).