Heme site

The very fast change relates to direct reduction of the Fe(III) center by the radical. The amount of absorbance change of this compared to the slow change can be understood if the hydrophobic nature of the heme site is considered. The rate of the slow change is similar for all systems since it involves a (common) intramolecular electron transfer. See (5.8.4). 

Exposure of ferri-heme-hemopexin to imidazole or KCN can displace one or both of the heme coordinating His residues, but millimolar concentrations are required (138). Other potential ligands such as azide or fluoride are inactive. This coordination stability of the ferri-heme-hemopexin bis-histidyl complex, despite the exposed heme site, is home out by thermal imfolding studies (Section IV,F). Reduction of the heme-hemopexin complex, however, has dramatic effects on its stability. 

Table III also shows the values of the equilibrium constants, KVAp for the conversion of iron nitrosyl complexes into the corresponding nitro derivatives. Keq decreases downwards, meaning that the conversions are obtained at a lower pH for the complexes at the top of the table. Thus, NP can be fully converted into the nitro complex only at pHs greater than 10. The NO+ N02 conversion, together with the release of N02 from the coordination sphere, are key features in some enzymatic reactions leading to oxidation of nitrogen hydrides to nitrite (14). The above conversion and release must occur under physiological conditions with the hydroxylaminoreductase enzyme (HAO), in which the substrate is seemingly oxidized through two electron paths involving HNO and NO+ as intermediates. Evidently, the mechanistic requirements are closely related to the structure of the heme sites in HAO (69). No direct evidence of bound nitrite intermediates has been reported, however, and this was also the case for the reductive nitrosylation processes associated with ferri-heme chemistry (Fig. 4) (25).

Unlike cytochrome P-450 reductase, NOS is a self-sufficient enzyme in that the oxygenation of its substrate, L-arginine, occurs at the heme-site in the N-terminal portion, termed the oxygenase domain, of the protein. Stoichiometric amounts of heme are present in NOS and are required for catalytic activity. 

A great role in substantiating the importance of electron tunneling reactions was played by the work of De Vault and Chance in 1966 where the characteristic time, t1/z, of electron transfer from the heme site of the cytochrome molecule to the chlorophyll molecule in a bacterium was shown to be constant within the temperature range of 130 to 4.2 K [4]. The temperature independence of t1/2 permitted one to reject a diffusion mechanism for the process. However, it was still impossible to exclude the possibility of the reaction to proceeding via direct contact between the active sites of the reacting molecules. 

For epoxidations, electron-rich alkenes react more readily, in case of p-substituted styrenes, with a concomitant increase in the amount of aldehyde by-product, suggesting formation of an unsymmetric intermediate. However, cw-stilbene is converted to cis-stilbene oxide in 82% yield, and the intermediate, if any, must be short-lived. Under the same conditions, the reaction of trans-stilbene is slow. A close parallel approach of the double bond to the active heme site seems to be essential for epoxidation. 

The roles of cytochromes b563717 and b2 in electron transfer have been investigated. Cytochrome b2 contains heme and flavin centres, and transfers electrons to cytochrome c after abstracting hydrogen from the substrate. Cytochrome c can withdraw an electron from the reduced three-electron donor flavo-cytochrome b2 only at the heme site. The donor heme site is then supplied with an electron from the flavin site, so that two further molecules of cytochrome c may ultimately be reduced.718... 

This electron transport and proton flow is controlled by at least three other proteins on the cytosolic side of the membrane, P47, P67, and P21rac (Figure 5). Note that this is a much simpler enzyme complex than the complex III (cyt bc ) of mitochondria which drives proton export from the interior of mitochondria based on quinol oxidation. Despite its simplicity, the neutrophil enzyme may have similarities to the complex three, since it essentially carries out an oxidation of a protonated two electron flavin by a nonprotonated cytochrome b complex with two heme sites. This is the essence of the mitochondrial enzyme in that the two electron quinol is oxidized by a cytochrome b with two heme components. 

The potentiometric observations in the presence of azide are as expected, if azide binds only to the oxidized low potential component (137). Here, too, the ligand appears to bind to the wrong component. If there is an initial difference in reducibility between the two hemes, the high potential (more readily reducible) component would be expected to have the lowest electron density and hence represent the binding site of preference for the (T-bonding ligand azide. If azide does bind to the high potential site, the potentiometric observations are difBcult to rationalize. Thus here, too, the assumption of an initial or inherent difference in the electron affinities of the two heme sites leads to difficulty in the completely rational interpretation of the experimental observations. 

On balance there seem to be fewer problems in interpretation if the two heme sites of oxidase(IV) or oxidase(O) are placed at the. same in-... 

---