Molybdenum iron protein cofactor

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. 

These clusters are each composed of eight iron atoms and seven sulfide ions. In the reduced form, each cluster takes the form of two 4Fe-3S partial cubes linked by a central sulfide ion. Each cluster is linked to the protein through six cysteinate residues. Electrons flow from the P cluster to the FeMo cofactor, a very unusual redox center. Because molybdenum is present in this cluster, the nitrogenase component is also called the molybdenum-iron protein (MoFe protein). The FeMo cofactor consists of two M-3Fe-3S clusters, in which molybdenum occupies the M site in one cluster and iron occupies it in the other. The two clusters are joined by three sulfide ions. The FeMo cofactor is also coordinated to a homocitrate moiety and to the a subunit through one histidine residue and one cysteinate residue. This cofactor is distinct from the molybdenum-containing cofactor found in sulfite oxidase and apparently all other molybdenum-containing enzymes except nitrogenase. 

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.)...

Fig. 1. Schematic illustration of the enzyme nitrogenase being composed of the molybdenum-iron (MoFe) protein, an oc2p2 tetramer with two unique iron-sulfur clusters (P-cluster) and two iron-molybdenum cofactors (FeMoco), and the iron protein with one [4Fe-4S]-cluster and two ATP binding sites.

Nitrogenase, which catalyzes the reduction of N2 to two molecules of NH3, has a different molybdenum -iron cofactor (FeMo-co). It can be obtained by acid denaturation of the very oxygen-labile iron-molybdenum protein of nitrogenase followed by extraction with d i methyl formamide.655,656 The coenzyme is a complex Fe-S-Mo cluster also containing homocitrate with a composition MoFe7S9-homocitrate (see Fig. 24-3). Nitrogenase and this coenzyme are considered further in Chapter 24. 

A number of molybdenum-iron-sulfide clusters have been synthesized as models for the cofactor.1478,1479,1486 At present there is no synthetic model which reproduces all of the spectroscopic properties of the cofactor. The ultimate test of a synthetic cluster would be its incorporation into the MoFe protein. Some clusters are shown in Figure 73. 

A cofactor can be extracted from the iron-molybdenum protein, using Af-methylformamide. This cofactor (called FeMoCo) has many spectroscopic properties in common with the native protein, especially the EXAFS spectrum, and activates the inactive large protein derived from Azobacter vinelandii UW45 mutant which cannot incorporate molybdenum. The cofactor contains no protein or peptide, but does contain molybdenum, iron, and sulfur in atomic ratios of 1 6-8 4-9. It is believed to contain the dinitrogen-binding site (presumably molybdenum) but there is no definitive proof of this. 

Cobalt B Enzymes Coenzymes Cytochrome Oxidase Iron Heme Proteins Electron Transport Iron Proteins with Dinuclear Active Sites Iron Proteins with Mononuclear Active Sites Iron-Sulfur Models of Protein Active Sites Metallocenter Biosynthesis Assembly. Metalloregulation Molybdenum MPT-containing Enzymes Nickel Enzymes Cofactors Nitrogenase Catalysis Assembly Photosynthesis Tungsten Proteins Vanadium in Biology Zinc DNA-binding Proteins. 

The best-characterized molybdenum nitrogenase comprises two metallosulfur proteins, that is, the molybdenum-iron (MoFe) protein and the iron (Fe) protein, both of which are essential for the enzymatic activity. The Fe protein is an U2 homodimer (encoded by nifH) of Mr 60 kDa. The two subunits are bridged by a [4Fe-4S] cluster and each has a MgATP binding site. The MoFe protein is an U2P2 tetramer (encoded by niJD and nifK) of Mr 220 kDa. It contains the [8Fe-7S] cluster (P cluster) that is bridged between each ap subunit pair and the [Mo-7Fe-9S-homocitrate] cluster (FeMo cofactor or FeMoco) that is located within each a snbunit. 

The enzyme system responsible for N2 reduction, called the nitrogenase complex, consists of two separate proteins. As outlined in Figure 20.4, one protein-called component I, nitrogenase, or molybdenumiron protein-catalyzes the reduction of N2, and the other-called component II, nitrogenase reductase, or iron protein-transfers electrons from ferredoxin or flavodoxin to component I. Both component I and component II contain Fe4S4 iron-sulfur clusters, and component I also contains molybdenum, in the form of a tightly bound iron-molybdenum cofactor (FeMoCo). 

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