Cytochrome hydroxylation mechanism

B3b system, although it certainly is not ruled out for the M. capsulatus (Bath) enzyme. In comparison to the cytochrome P-450 system, the hydroxylation mechanism for both MMO systems either has a rebound rate constant which is much larger and/or it takes place by an alternative pathway to classical radical rebound. 

In addition to a well-known NADPH-dependent hydroxylation mechanism (Reaction (2)), cytochrome P-450 is able to catalyze the oxidation of substrates by peroxygenase mechanism (Reaction (8)) where XOOH presents the peroxy compound acting as the oxygen donor. 

The most ambiguous point in the oxynoid hydroxylation mechanism with cytochrome P-450 is the question about the formation and origin of the oxygen intermediate Fe3+ O, where the oxygen atom (possessing six electrons in the external cover) is coordinated withFe3+. 

Studies with cell-free hydroxylases suggest that the hydroxylation mechanisms are complex. It is assumed that an electron transport system involving an NADPH-dependent flavoprotein, an iron-sulfur protein, and cytochrome P-450 is involved. In the case of the steroid 15/S-hydroxylase system of Bacillus megaierium, these three components have been demonstrated15. The 1 la-hydroxylase of Rhizopus nigricans is also an enzyme of the P-450 monooxygenase type which works with an NADPH-cytochrome P-450 reductase. In this case the enzyme complex is associated with the endoplasmic reticulum of the mycelial cells34. 

Figure 26.26. Cytochrome P450 Mechanism. These enzyme-bind O2 and use one oxygen atom to hydroxylate their substrates.

Mechanistical studies, for example, on the stereochemistry of the I -hydroxylation of (R)- and (S)-r-deuterated-phenylelhane by purified cytochrome P450lm2 have demonstrated that the product stereoselectivity most probably results from constraints of the substrate binding site provided by the protein environment, rather than the intrinsic hydroxylation mechanism of cytochrome P450 (White et al., 1986). 

In users of Ge preparations, particularly high accumulations of this element were found in spleen, renal cortex, brain, and skeletal muscle (Nagata et al. 1985). Organic Ge preparations are biotransformed by the liver microsomal cytochrome P-450 enzyme system, probably through a C-hydroxylation mechanism (Prough et al. 

Table 10.2. Hydroxylation Mechanisms Catalyzed by Cytochrome P450...

Kumar D, de Visser SP, Sharma PK, Cohen S, Shaik S (2004) Radical clock substrates, their C-H hydroxylation mechanism by cytochrome P450, other reactivity patterns what does theory reveal about the clocks behavior J Am Chem Soc 126 1907-1920... 

C. Hydroxylation Mechanism of ML-236B to Pravastatin by a Novel Actinomyecte Cytochrome P-4S0... 

As a class of compounds, the two main toxicity concerns for nitriles are acute lethality and osteolathyrsm. A comprehensive review of the toxicity of nitriles, including detailed discussion of biochemical mechanisms of toxicity and stmcture-activity relationships, is available (12). Nitriles vary broadly in their abiUty to cause acute lethaUty and subde differences in stmcture can greatly affect toxic potency. The biochemical basis of their acute toxicity is related to their metaboHsm in the body. Following exposure and absorption, nitriles are metabolized by cytochrome p450 enzymes in the Hver. The metaboHsm involves initial hydrogen abstraction resulting in the formation of a carbon radical, followed by hydroxylation of the carbon radical. MetaboHsm at the carbon atom adjacent (alpha) to the cyano group would yield a cyanohydrin metaboHte, which decomposes readily in the body to produce cyanide. Hydroxylation at other carbon positions in the nitrile does not result in cyanide release. 

Cytochrome P450s catalyze reactions that introduce one atom of oxygen derived from molecular oxygen into the substrate, yielding a hydroxylated product. NADPH and NADPH-cytochrome P450 reductase are involved in the complex reaction mechanism. 

The mechanism of the cytochrome P450i4D]y conversion of a C14 methyl sterol into formate and the was studied using 02, and it could be shown that the hydroxyl oxygen atom in the formate that was produced contained one atom of (Shyadehi et al. 1996). 

Ingelman-Sundberg, M. and Ekstrom, G. (1982). Aniline is hydroxylated by the cytochrome P-450-dependent hydroxyl radical-mediated oxygenation mechanism. Biochem. Biophys. Res. Commun. 106, 625-631. 

Colquhoun and Schumacher [98] have shown that y-linolcnic acid and eicosapentaenoic acid, which inhibit Walker tumor growth in vivo, decreased proliferation and apoptotic index in these cells. Development of apoptosis was characterized by the enhancement of the formation of reactive oxygen species and products of lipid peroxidation and was accompanied by a decrease in the activities of mitochondrial complexes I, III, and IV, and the release of cytochrome c and caspase 3-like activation of DNA fragmentation. Earlier, a similar apoptotic mechanism of antitumor activity has been shown for the flavonoid quercetin [99], Kamp et al. [100] suggested that the asbestos-induced apoptosis in alveolar epithelial cells was mediated by iron-derived oxygen species, although authors did not hypothesize about the nature of these species (hydroxyl radicals, hydrogen peroxide, or iron complexes ). 

FIGURE 24.1 Mechanism of hydroxylation and reduction of substrates by cytochrome P-450. (From TD Porter, MJ Coon. J Biol Chem 266 13469-13472, 1991. With permission.)... 

If the mechanism of superoxide production in microsomes by NADPH-cytochrome P-450 reductase, NADH-cytochrome b5 reductase, and cytochrome P-450 is well documented, it cannot be said about microsomal hydroxyl radical production. There are numerous studies, which suggest the formation of hydroxyl radicals in various mitochondrial preparations and by isolated microsomal enzymes. It has been shown that the addition of iron complexes to microsomes stimulated the formation of hydroxyl radicals supposedly via the Fenton... 

Although the oxidation (hydroxylation) of hydrocarbons is usually believed to occur via hydrogen atom abstraction [51], the one-electron transfer mechanism of cytochrome P-450 catalyzed oxidation has also been proposed for the oxidation of Ar, Ar-dialkylaniIines [52]. This mechanism (Figure 24.4) is generally preferred for the substrates with low reduction... 

Similar to some other antioxidants, pyrrolopyrimidines do not contain active free radical scavenging groups such as phenolic or thiolic substituents. At present, at least two different mechanisms of their antioxidant activity have been proposed [307], It was suggested that pyrrolopyrimidines, which are electron donating compounds, can be oxidized by hydroxyl or peroxyl radicals or hydroxylated by cytochrome P-450 forming phenolic metabolite... 

Adults require 1-2 mg of copper per day, and eliminate excess copper in bile and feces. Most plasma copper is present in ceruloplasmin. In Wilson s disease, the diminished availability of ceruloplasmin interferes with the function of enzymes that rely on ceruloplasmin as a copper donor (e.g. cytochrome oxidase, tyrosinase and superoxide dismutase). In addition, loss of copper-binding capacity in the serum leads to copper deposition in liver, brain and other organs, resulting in tissue damage. The mechanisms of toxicity are not fully understood, but may involve the formation of hydroxyl radicals via the Fenton reaction, which, in turn initiates a cascade of cellular cytotoxic events, including mitochondrial dysfunction, lipid peroxidation, disruption of calcium ion homeostasis, and cell death. 

Although pathway 2 in the oxidation process (Scheme 2) may be considered analogous to mechanisms proposed for carbon hydroxylations catalyzed by cytochrome P-450, abstraction of an electron from the lone pair on nitrogen (pathway 1) would be a more likely first step in these types of reactions. It is reasonable to assume that the nature of substituents R, R2, and R3 would greatly influence the rate and path of reaction. The mechanistic possibilities in Scheme 2 are undoubtedly simplistic in their representation of the active oxygen species of cytochrome P-450 and are by no means comprehensive. However, these pathways do serve to illustrate.the role of radical substrate intermediates in cytochrome P-450-catalyzed reactions. More detailed analyses of mechanistic studies on these and other cytochrome P-450-mediated reactions can be found in recent reviews on the subject 49, 50, 60). 

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