Molybdenum iron protein cofactor structure

Bolen, J.T., Cambasso, N., Muchmore, S.W., Morgan, T.V., and Mortenson, L. E. (1993) Structure and Environment of metal clusters of the nitrogenase molybdenum iron protein from Clostridium pasterianum, in Stiefel, E.I., Coucouvanis, D., and ewton, W.E. (eds.), Molibdenum Enzymes, Cofactors, and Model Systems, Am. Chem. Soc., Wahington, DC. 

The sequential electron transfer path in nitrogenase is followed first models of the Fe4S4 cluster of the iron-protein are discussed, then mimics of the P-cluster in the molybdenum-iron protein, and finally structural and functional models of the FeMo-cofactor are summarized. 

VU Fig. 28.18 The structures of the two types of cluster unit present in the nitrogenase molybdenum-iron protein isolated from Azotobacter vinelandii (a) the P-cluster in its reduced state and (b) the FeMo cofactor. Colour code Fe, green Mo, pale grey S, yellow C, grey N, blue O, red. Each non-terminated stick represents the connection of a coordinated amino acid to the protein backbone. 

T. V. Morgan and L. E. Mortenson, Structure and environment of metal clusters in the nitrogenase molybdenum-iron protein from Clostridium pasteurianum, in Molybdenum Enzymes, Cofactors and Model Systems , eds. E. I. Stiefel,... 

Figure 4.2. Schematic models of Fe-Mo cofactor with possible binding of dinitrogen substrate (a) and P cluster (b). Y indicates a bridging ligand. (Part a adapted, with permission, from Kim, J., and Rees, D. C. Structural models for metal centres in the nitrogenase molybdenum-iron protein. Science 257 1677-82. Copyright 1992, American Association for the Advancement of Science. Part b adapted, with permission, from Chan, M. K., Kim, J., and Rees, D. C. The nitrogenase Fe-Mo-cofactor and P-cluster pair 2.2A resolution structures. Science 260 792-4. Copyright 1993, American Association for the Advancement of Science.)...

FeMoco can be extracted from the MoFe protein into A(-methylfor-mamide (NMF) solution 32) and has been analyzed extensively using a wide range of spectroscopic techniques both bound to the protein and in solution after extraction from it (33). The extracted FeMoco can be combined with the MoFe protein polypeptides, isolated from strains unable to synthesize the cofactor, to generate active protein. The structure of the FeMoco is now agreed 4, 5, 7) as MoFeTSg homocitrate as in Fig. 4. FeMoco is bound to the a subunit through residues Cys 275, to the terminal tetrahedral iron atom, and His 442 to the molybdenum atom (residue numbers refer to A. vinelandii). A number of other residues in its environment are hydrogen bonded to FeMoco and are essential to its activity (see Section V,E,2). The metal... 

Synthetic structural models for clusters within the nitrogenase proteins or of the molybdenum or vanadium iron - sulphur cofactors isolated from them, are at an eaily stage of development and there are as yet no well - defined systems which model electron - transfer chemistry involving the binding and reduction of molecular nitrogen or other substrates dinitrogen binding to synthetic iron - sulphur clusters or heterometallic iron - sulphur clusters of any type has not been established structurally nor spectroscopically, [44- 46]. 

One type of the constituent metallocenters in the MoFe protein has the properties of a somewhat independent structural entity. This component, referred to as the FeMo cofactor (FeMo-co), was first identified by Shah and Brill (1977) as the stable metallocluster extracted from acid-denatured MoFe protein. The FeMo-co was able to fully activate a defective protein in the extracts of mutant strain UW45, a protein which subsequently was shown to contain the P clusters but not the EPR-active center. The isolated cofactor accounted for the total S = t system observed by EPR and Mdssbauer spectroscopies of the holo-MoFe protein (Rawlings et al., 1978). Elemental analysis indicated a composition of Mo Fee-8 Se-g for the cofactor, which, if there are two FeMo-co s per a2 2> accounts for all the molybdenum and approximately half the iron in active enzyme (Nelson etai, 1983). Although FeMo-co has been extensively studied [reviewed in Burgess (1990)] the structure remains enigmatic. To date, all attempts to crystallize the cofactor have failed. This is possibly due to the instability and resultant heterogeneity of the cofactor when removed from the protein. Also, there is a paucity of appropriate models for spectral comparison (see Coucouvanis, 1991, for a recent discussion). Final resolution of this elusive structure may require its determination as a component of the holoprotein. 

FeMoco, both as a constituent of the FeMo protein and an isolated entity, has been the subject of detailed spectroscopic examination. 57Fe Mossbauer and EPR studies of the cofactor have been interpreted in terms of an S = centre that contains one molybdenum and ca. six irons in a spin-coupled structure. The EPR signal serves as a valuable fingerprint of FeMoco furthermore, release of FeMoco from the FeMo protein produces an EPR spectrum with broader features, but the same profile, thereby indicating that the core of this cluster is little changed by the extraction procedure. Treatment of FeMoco with ca. one equivalent of... 

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