Structure of nitrogenase

Figure 5 Some metallo-enzymes are formed by a set of subunits. Shown here is the structure of nitrogenase that has an energized supply of electrons (energy from ATP) via two iron/sulfur clusters and an active site FeMoco shown in the inset. There are two distinct protein units linked to one supply line of a separate unit. Note there is a light non-metal in a hole in the structure of Fe7MoS9 but, as it is of uncertain nature, it is not shown.

Garcia-Horsman JA, Berry E, Shapleigh JP, Alben JO, Gennis RB (1994) A novel cytochrome c oxidase from Rhodobacter sphaeroides that lacks Cua. Biochemistry 33 3113-3119 Georgiadis MM, Komiya H, Chakrabarti P, Woo D, Komuc JJ, Rees DC (1992) Crystallographic structure of nitrogenase iron protein from Azotobacter vinelandii. Science 257 1653-1659... 

Crystal structures of nitrogenase and nitrogenase reductase (Figure 20.5) are now available and may help scientists understand how the enzyme system for nitrogen fixation operates. 

Orme-Johnson, W. Nitrogenase Structure Where to Now Science 257, 1639-1640 (1992). [Thoughts about nitrogen fixation based on the determination of the structure of nitrogenase by X-ray crystallography]. 

A famous example is the structure of Nitrogenase MoFe-Protein, a protein that contains a Fe7MoS9 cluster. The inside of this cluster is about 4 A wide with six iron atoms closest to the centre, and older crystal stmctures had been determined at resolutions of about 2 A. Termination of the Fourier summation at that resolution creates an artefactual miiumum in the electron density of about —0.2 electrons about 2 A away from each iron atom. These spurious minima from all heavy atoms in the... 

In this section DFT treatments of the iron-molybdenum cofactor and the activation, reduction and protonation of N2 proceeding at this cluster are presented. The earliest of these calculations appeared after publication of the first crystal structures of nitrogenase, and this was well before the discovery of the central atom X. After the discovery of X and its identification as carbon these treatments were, in part, updated. Here we will focus both on the basic theoretical methods to treat the electronic structure of the FeMoco and the reduction of N2 mediated by this cluster. 

Schindeiin N, Kisker C, Sehiessman J L, Howard J B and Rees D C 1997 Structure of ADP center dot AiF(4)(-)-stabiiized nitrogenase compiex and its impiications for signai transduction Nature 387 370-6... 

Fe—M—S complexes derived from MS4 anions (M=Mo, W) and their possible relevance as analogues for structural features in the Mo site of nitrogenase. D. Coucouvanis, Acc. Chem. Res., 1981,... 

In late 1992 the first crystal structures of the Fe and MoFe proteins of Mo nitrogenase frora Azotobacter vinelandii were published (1-3). 

These were relatively low-resolution structures, and with refinement some errors in the initial structural assignments have been detected (4-7). Since the structures were first reported the subject has been extensively reviewed in this series (8) and elsewhere 9-15). This review will focus on the structure, biosynthesis, and function of the met-allosulfur clusters found in nitrogenases. This will require a broader overview of some functional aspects, particularly the involvement of MgATP in the enzymic reaction, and also some reference will be made to the extensive literature (9, 15) on biomimetic chemistry that has helped to illuminate possible modes of nitrogenase function, although a detailed review of this chemistry will not be attempted here. This review cannot be fully comprehensive in the space available, but concentrates on recent advances and attempts to describe the current level of our understanding. 

However, when the X-ray crystal structure of the MoFe protein was examined, it was clear that homocitrate could not directly hydrogen bond to the histidine, since the carboxylate group and imidazole are stacked parallel to each other in the crystal. Nevertheless, as noted in the previous section, studies on model complexes have suggested that homocitrate can become monodentate during nitrogenase turnover, with the molybdenum carboxylate bond breaking to open up a vacant site at molybdenum suitable for binding N2. 

A preparation of the third nitrogenase from A. vinelandii, isolated from a molybdenum-tolerant strain but lacking the structural genes for the molybdenum and vanadium nitrogenases, was discovered to contain FeMoco 194). The 8 subunit encoded by anfG was identified in this preparation, which contained 24 Fe atoms and 1 Mo atom per mol. EPR spectroscopy and extraction of the cofactor identified it as FeMoco. The hybrid enzyme could reduce N2 to ammonia and reduced acetylene to ethylene and ethane. The rate of formation of ethane was nonlinear and the ethane ethylene ratio was strongly dependent on the ratio of nitrogenase components. 

The elucidation of the crystal structures of two high-spin EPR proteins has shown that the proposals for novel Fe-S clusters are not without substance. Two, rather than one novel Fe-S cluster, were shown to be present in nitrogenase, the key enzyme in the biotic fixation of molecular nitrogen 4, 5). Thus the FeMoco-cofactor comprises two metal clusters of composition [4Fe-3S] and [lMo-3Fe-3S] bridged by three inorganic sulfur atoms, and this is some 14 A distant from the P-cluster, which is essentially two [4Fe-4S] cubane moieties sharing a corner. The elucidation of the crystal structure of the Fepr protein (6) provides the second example of a high-spin EPR protein that contains yet another unprecedented Fe-S cluster. 

Structure and Function of Nitrogenase Douglas C. Rees, Michael K. Chan, and Jongsun Kim... 

Researchers are working to understand how this enzyme works. Research on nitrogenase takes two main forms. One is an examination of the structure and operation of the enzyme to determine the details of the reactions by which N2 is converted to ammonia. The other form is the s Tithesis of artificial catalysts that mimic the operation of nitrogenase. 

Figure 2.10 Schematic structures of (a) sulfite reductase of Escherichia coli in which a 4Fe-4S cluster is linked via a cysteine to the iron in a sirohaem (b) P cluster of nitrogenase (c) FeMoCo cluster of nitrogenase (d) the binuclear site in Desulforibrio gigas hydrogenase.

Figure 1 Structure of the MoFe7SgN cluster of FeMo-co in Mo nitrogenase...

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