Catalytic cycle of CYP

In the catalytic cycle of CYP, reducing equivalents are transferred from NADPH to CYP by a flavoprotein enzyme known as NADPH-cytochrome P450 reductase. The evidence that this enzyme is involved in CYP monooxygenations was originally derived from the observation that cytochrome c, which can function as an artificial electron acceptor for the enzyme, is an inhibitor of such oxidations. This reductase is an essential component in CYP-catalyzed enzyme systems reconstituted from purified components. Moreover antibodies prepared from purified reductase are inhibitors of microsomal... 

FIGURE 9.R.1 The catalytic cycle of CYP 450 during the hydroxylation of an alkane (R—H), in an overall process which replaces the C—H bond with a C—OH bond. Except for the resting state, 1, where the porphyrin ring is drawn explicitly, for all the subsequent species, the ring is represented by two heavy bars flanking Fe. All the intermediate species are labeled with bold numerals. The active species, which breaks the C—H bond, is indicated as 7. 

Figure 4.2 The catalytic cycle of the cytochrome(s) P-450 monooxygenase (MFO) system. aFor explanation of step 4, see text. Abbreviation Cyp, cytochromes P-450 MFO, mixed function oxidase. Source From Ref. 1.

Several unique features of the catalytic cycle of the FMOs are important for understanding the mechanism by which they oxidize xenobiotics. The catalytic mechanism for the FMO has been shown to involve the formation of an enzyme bound 4a-hydroperoxyl-flavin (Figure 10.6) in an NAD PH and 02 dependent reaction. Reduction of the flavin by NAD PH occurs before binding of oxygen can occur, and activation of oxygen by the enzyme occurs in the absence of substrate by oxidizing NADPH to form NADP and peroxide. Finally, addition of the substrate to the peroxyflavin complex is the last step prior to oxygenation. This is in contrast to the CYP catalytic cycle in which the substrate binds to the oxidized enzyme which is subsequently reduced. 

FIGURE 33.2 Catalytic cycle of cytochrome P450 (CYP) monooxygenase. 

H202 may be produced as a by-product in the catalytic cycle of the CYP enzymes described later. Diafenthiuron therefore becomes more active in sunshine, and piperonyl butoxide that inhibits CYP enzymes makes diafenthiuron less toxic. However, some CYP enzymes are also important in the detoxication of diafenthiuron, as shown in Figure 3.4. Diafenthiuron may... 

Figure 8.8 The catalytic cycle of the CYP enzymes. The substrate binds to a hydro-phobic site of the enzyme (I —> II). This leads to a shift in the valence electrons of iron, from low-spin to high-spin status, and an uptake of an electron from cytochrome P450 reductase (II —> III). Oxygen is then added (III —> IV) to the iron and another electron is added (IV V). Some of the intermediates may lose reactive oxygen species (IV —> II), which has harmful consequences for the cell. There are some electronic rearrangements (V —> VI —> VII). The product (RO) leaves the enzyme and its ground state is restored (VII —> I). RO is often more toxic than the substrate (R) and may be an alcohol, a phenol, or an epoxide that can be rendered harmless by other enzymes and made ready for excretion.

Figure 9.1 CYP catalytic cycle. The sequential two-electron reduction of CYP and the various transient intermediates were first described in the late 1960s [206], The sequence of events that make up the CYP catalytic cycle is shown. The simplified CYP cycle begins with heme iron in the ferric state. In step (i), the substrate (R—H) binds to the enzyme, somewhere nearthe distal region of the heme group and disrupts the water lattice within the enzymes active site [207], The loss of water elicits a change in the heme iron spin state (from low spin to high spin) [208]. Step (ii) involves the transfers of an electron from NADPH via the accessory flavoprotein NADPH-CYP reductase, with the electron flow going from the reductase prosthetic group FAD to FMN to the CYP enzyme [206,209]. The...

Figure 30. Natural catalytic cycle for the hydroxylation of organic compounds catalyzed by the Cytochrome (Cyt) P-450cam monooxygenase system (Cyp = Cyt. P-450 hemeprotein Pdx = putidaredoxin).

Gunsalus laboratory, the P450(, system allowed biochemical and biophysical investigation of the CYP catalytic cycle as well as of the genetics of a bacterial catabolic plasmid. This typical CYP system was found to require a ferredoxin and ferredoxin reductase for catalytic activity, unlike the model for eukaryote CYPs, CYP102A1 or P450g 3, which was discovered in the Fulco laboratory and consisted of a fusion polypeptide containing CYP and reductase domains. ... 

The broad application of cytochrome P450 monooxygenases for synthetic purposes is still hampered by their comparably low catalytic rates and their usually rather limited stability, especially in isolated form, because of the possible formation of reactive oxygen species during the catalytic cycle (Scheme 36.17). Nevertheless, industrial applications of CYPs in the syntheses of certain drugs, e.g., the stereoselective hydroxylation of steroids or the oxidation of compactin 43 to pravastatin 44, a potent cholesterol-lowering agent, have already been reported (Scheme 36.18). ... 

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