Nitrogenase substrate binding site

Although FeMo-cofactor is clearly knpHcated in substrate reduction cataly2ed by the Mo-nitrogenase, efforts to reduce substrates using the isolated FeMo-cofactor have been mosdy equivocal. Thus the FeMo-cofactor s polypeptide environment must play a critical role in substrate binding and reduction. Also, the different spectroscopic features of protein-bound vs isolated FeMo-cofactor clearly indicate a role for the polypeptide in electronically fine-tuning the substrate-reduction site. Site-directed amino acid substitution studies have been used to probe the possible effects of FeMo-cofactor s polypeptide environment on substrate reduction (163—169). Catalytic and spectroscopic consequences of such substitutions should provide information concerning the specific functions of individual amino acids located within the FeMo-cofactor environment (95,122,149). 

Early data on the substrate and inhibitor reactions of nitrogenase were interpreted in terms of five binding sites, with competitive, noncompetitive, unclassified, and negative inhibition being observed (127). This apparent complexity can be readily rationalized in terms of the Lowe—Thorneley scheme (Fig. 9) by assuming that different substrates bind at different oxidation states of the same site. 

Cluster models of the polymetallic aggregates in nitrogenase should (1) contain structural features akin to those defined by the crystal structure (2) possess or approximate the observed stoichiometry (3) provide coordination sites at which substrate binding and activation may occur. A model need not mimic protein-bound cofactor reactivity, since isolated cofactors do not reduce sub-... 

While Fe(SCys)4, [2Fe-2S], [3Fe-4S] and [4Fe-4S] clusters all function as one-electron donors or acceptors, the more complex double-cubane [8Fe-7S] cluster that is found only in nitrogenases (see Nitrogenase Catalysis Assembly) has the potential to mediate two-electron transfer processes.Three methods have been employed to functionalize Fe-S centers for substrate binding and activation. The first involves having an accessible Fe coordination site as in the mononuclear Fe centers of nitrile hydratase and SOR, and the [4Fe-4S] clusters at the active sites of hydratases/dehydratases and radical-5-adenosylmethionine (SAM) enzymes.Indeed the recent recognition of the importance of the superfamily of radical-SAM enzymes in initiating radical reactions, via cluster-mediated reductive cleavage of SAM to yield a... 

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