Binuclear iron centres

In analogy with peroxidase/P-450 mechanisms it is possible to devise a mechanism for these reactions that involve ferryl intermediates (Fig. 5). In favour of these analogies is the fact that H2O2 will act as an electron and oxygen donor to the di-ferric form of both enzymes, bypassing the need for a reductant and molecular oxygen [73-75]. However, it must be stressed that these mechanisms are still controversial [76]. Although the compound II-equivalent intermediate 

Mechanisms for those enzymes acting as monooxygenases, e.g. mammalian tyrosine hydroxylase [87], Pseudomonas oleovorans monooxygenase [88] or 4-methoxybenzoate monooxygenase [84] can be devised that are essentially identical to those of cytochrome P-450 (Fig. 4) in this case the transient compound I-type intermediate is formalised as Fev=0, as there is no porphyrin ring to accept a cation radical. However, alternative mechanisms have been suggested that involve a direct attack on the substrate by the peroxy intermediate, Fe3+-C 2 [78,84], 

Methods for detecting ferryl iron and protein free radicals 4.1. X-ray crystallography 

Several of the proteins with ferryl intermediates have been crystalised at sufficient resolution to allow the elucidation of their 3-dimensional structure. These include cytochrome c peroxidase [95], horseradish peroxidase [96], catalase [97], myeloperoxidase [98], ribonucleotide reductase [99], cytochrome P-450 [100] and myoglobin [101]. Of these only cytochrome c peroxidase has proved stable enough to crystallise with the iron in the ferryl form [26]. High-resolution structures exist for small FeIV model compounds, both in the presence [102] and absence [7,8] of an Fe=0 bond. These compounds can have sulphur, nitrogen and chloride ligation to the iron and the iron can be five [7,8] or six [8] coordinate. 

Compound I crystals of cytochrome c peroxidase can be generated by the direct addition of H2O2 to crystals of the ferric complex [7]. These crystals are stable for four hours at — 15°C, which with the improvements in data collection afforded by area detectors [103] allows a difference crystal structure to be determined relative to that of the ferric enzyme. Unfortunately the resolution 

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Dobias B (1984) Surfactant Adsorption on Minerals Related to Flotation. 56 91-147 Doi K, Antanaitis BC, Aisen P (1988) The Binuclear Iron Centres of Uteroferrin and the Purple Acid Phosphatases. 70 1-26 Domcke W, see Bradshaw AM (1975) 24 133-170 Dophin D, see Morgan B (1987) 64 115-204... 

Figure 55 Structures for the binuclear iron centre in hemerythrin...

Ribonucleotide reductase catalyses the reduction of the four common ribonucleotides to their corresponding deoxyribonucleotides, an essential step in DNA synthesis. All four ribonucleotides are reduced by the same enzyme [77], The enzyme (250 000 mol. wt.) is a complex of two proteins Mi which contains substrate and redox-active sulphydryl groups and M2 which contains both a (x-oxo-bridged binuclear iron centre (Fig. 5) [77] and a tyrosine moiety sidechain which exists as a free radical stabilised by the iron centre [78], This radical, which is only 5.3 A away from iron centre 1, has access to the substrate-binding pocket and is essential for enzyme activity. Electrons for the reduction reaction are supplied from NADPH via thioredoxin, a small redox-active protein. 

Oxo-bridged binuclear iron centres in oxygen transport, oxygen reduction and oxygenation have been studied and reviewed by Sanders-Loer and coworkers . Oxidations and reductions in general, including reported reductions and oxidations of biological interest, have been reviewed critically by Fleet . 

Tartrate-resistant acid phosphatase is characterised for activated macrophages (Efstratiadis and Moss 1985). Tartrate-resistant acid phosphatase contains a binuclear iron centre that has been shown to generate reactive oxygen species. Raisa-nen et al. (2001) showed that the murine macrophage-like cell line RAW-264 overexpressing tartrate-resistant acid phosphatase produced elevated levels of hydroxyl radicals compared to parental cells. 

Binuclear, Oxo-bridged Iron Centres and the Iron-tyrosinate Proteins... 

It seems probable that other redox centres contain this binuclear iron structure, but that this has not yet been recognized. For example, a non-heme iron protein of the methane monooxygenase from Methylococcus capsulatus (Bath), which functions as the oxygenase in equation (28), has been described as having a novel iron centre which is not an iron-sulfur cluster. This may well be an oxo-bridged system. Analysis suggests 2.3 Fe per molecule of protein. 

However, it is possible to detect a tyrosine radical optically in ribonucleotide reductase, as there is only a relatively weak competing absorption from the binuclear non-haem iron centre [164]. A distinct sharp peak is seen that is not present in proteins that have been treated with the radical scavenger hydroxyurea [165,166] nor is it present in proteins such as haemerythrin or methane monooxygenase, which have similar active-site structures, but lack... 

The Mossbauer spectrum of native ribonucleotide reductase is very similar to that of oxyhaemerythrin, both having a binuclear Fein-Fein cluster with 5 = 0 [127,188]. This suggests that there is negligible coupling of the iron centre to the distant (5A) tyrosine-122 radical in ribonucleotide reductase. This was confirmed by the fact that removing the tyrosine radical with hydroxyurea or hydroxylamine had no effect on the spectrum [188]. However, by rapid-freezing... 

Several diverse metal centres are involved in the catalysis of monooxygenation or hydroxylation reactions. The most important of these is cytochrome P-450, a hemoprotein with a cysteine residue as an axial ligand. Tyrosinase involves a coupled binuclear copper site, while dopamine jS-hydroxylase is also a copper protein but probably involves four binuclear copper sites, which are different from the tyrosinase sites. Putidamonooxin involves an iron-sulfur protein and a non-heme iron. In all cases a peroxo complex appears to be the active species. 

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