Oxidations

5.05.2.1 Cytochromes P450

5.05.2.1.1 Occurrence, multiplicity, catalytic cycle, oxygen activation, and selectivity

As components of a biological defense system the cytochrome P450 enzymes2 are by far the most dominant and important (Table 1). They embody a superfamily of monooxygenases that are found in living organisms, ranging from bacteria to humans.3 In humans and other mammals the cytochromes P450 are found in highest concentration in the endoplasmic reticulum of the liver, and in lower concentration in other tissues (e.g., brain, lung, kidney, and intestine). In addition to defense, some cytochromes P450 play a significant role in normal physiology. In the human, for example, specialized cytochromes P450 found in the adrenal cortex are involved in the syntheses of steroid hormones, while others are involved in the oxidative transformation of fatty acids and the production of prostaglandins. However, the focus of this chapter will be on the cytochromes P450 involved in drug metabolism. These specific cytochromes P450 are found primarily in three families,4 CYP1, CYP2, and CYP3 (see 5.06 Principles of Drug Metabolism 2: Hydrolysis and Conjugation Reactions; 5.07 Principles of Drug Metabolism 3: Enzymes and Tissues, for an extensive discussion of the biological and structural properties of the cytochromes P450). Of the cytochromes P450, CYP3A4 is, arguably, the most important, as it has been found to contribute significantly to the metabolism of approximately half of all drugs that have been administered to humans or are in current use.5

All the cytochromes P450 work by activating O2. In the cycle6'7 that describes the catalytic role, O2 is split into two oxygen atoms; one atom is reduced to water, while the second is transferred to the substrate (Figure 1).

In the resting state the charge of heme Fe3 + is counterbalanced by two negatively charged nitrogen atoms from the porphyrin ring plus a negatively charged cysteine thiolate group that anchors heme to the protein. Heme Fe3 + is primarily in the low-spin form characterized by hexacoordinated heme Fe3 + in which Fe3 + lies in the plane of the porphyrin ring. Four of the ligand sites are occupied by the four imidazole nitrogen atoms, and the fifth by cysteine thiolate. The sixth is presumed to be occupied by a molecule of water.

• Step 1 - the substrate, RH, associates with the active site of the enzyme, and perturbs the spin state equilibrium. Water is ejected from the active site, and the high-spin pentacoordinated substrate-bound form becomes dominant. Fe3 + is puckered out and above the plane in the direction of the sixth ligand site. The change in spin state alters the redox potential of the system. Substrate bound enzyme is now more easily reduced.

• Step 2 - NADPH-dependent cytochrome P450 reductase transfers an electron to heme Fe3 +.

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