Cytochrome b5 reductase

Hereditary methemoglobinemia is classified into three types a red blood cell type (type I), a generalized type (type II), and a blood cell type (type HI). Enzyme deficiency of type I is limited to red blood cells, and these patients show only the diffuse, persistent, slate-gray cyanosis not associated with cardiac or pulmonary disease. In type II, the enzyme deficiency occurs in all cells, and patients of this type have a severe neurological disorder with mental retardation that predisposes them to early death. Patients with type III show symptoms similar to those of patients with type I. The precise nature of type III is not clear, but decreased enzyme activity is observed in all cells (M9). It is considered that uncomplicated hereditary methemoglobinemia without neurological involvement arises from a defect limited to the soluble cytochrome b5 reductase and that a combined deficiency of both the cytosolic and the microsomal cytochrome b5 reductase occurs in subjects with mental retardation. Up to now, three missense mutations in type I and three missense mutations, two nonsense mutations, two in-frame 3-bp deletions, and one splicing mutation in type n have been identified (M3, M8, M31). 

T17. Tomatsu, S., Kobayashi, Y., Fukumaki, Y., Yubisui, T., Orii, T., and Sakaki, Y., The organization and the complete nucleotide sequence of the human NADH-cytochrome b5 reductase gene. Gene 80,353-361 (1989). 

Microsomal NADPH-Cytochrome P-450 Reductase and NADH cytochrome b5 Reductase... 

MICROSOMAL NADPH-CYTOCHROME P-450 REDUCTASE AND NADH CYTOCHROME b5 REDUCTASE... 

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... 

Qrunones can accept one or two electrons to form the semiquinone anion (Q ") and the hydroquinone dianion (Q ). Single-electron reduction of a quinone is catalyzed by flavoenzymes with relatively low substrate selectivity (Kappus, 1986), for instance NADPH cytochrome P-450 reductase (E.C. 1.6.2.3), NADPH cytochrome b5 reductase (E.C. 1.6.2.2), and NADPH ubiquinone oxidoreductase (E.C. 1.6.5.3). The rate of reduction depends on several interrelated chemical properties of a quinone, including the single-electron reduction potential, as well as the number, position, and chemical characteristics of the substituent(s). The flavoenzyme DT-diphorase (NAD(P)H quinone acceptor oxidoreductase E.C. 1.6.99.2) catalyzes the two-electron reduction of a quinone to a hydroquinone. 

Eight human brain tumors contained DT-diaphorase, NADH cytochrome b5 reductase and NADPH cytochrome P450 reductase as assessed by enzyme activity and WB (Rampling et al., 1994). 

In humans, a variety of HA derivatives that are obtained by bioactivation of arylamine drugs (i.e. A-hydroxylation) or of pro-drugs that contain the NH—OH function, and whose activation requires the reduction of the hydroxylamine nitrogen atom, have been shown to undergo enzymatic reduction by the cytochrome f)5/NADH cytochrome b5 reductase system. ... 

The endoplasmic reticulum is composed of a convoluted network of channels and so has a large surface area. Apart from cytochromes P-450, the endoplasmic reticulum has many enzymes and functions, besides the metabolism of foreign compounds. These include the synthesis of proteins and triglycerides and other aspects of lipid metabolism and fatty acid metabolism. Specific enzymes present on the endoplasmic reticulum include cholesterol esterase, azo reductase, glucuronosyl transferase, NADPH cytochromes P-450 reductase and NADH cytochrome b5 reductase and cytochrome b5. A FAD-containing monooxygenase is also found in the endoplasmic reticulum, and this is discussed later in this chapter. 

Step 4. Addition of the second electron from NADPH via P-450 reductase. Step 4. Alternatively, the electron may be donated by NADH via cytochrome b5 reductase and cytochrome bs. This alternative source of the second electron step is still controversial. 

Tertiary amine oxides and hydroxy la mines are also reduced by cytochromes P-450. Hydroxylamines, as well as being reduced by cytochromes P-450, are also reduced by a flavoprotein, which is part of a system, which requires NADH and includes NADH cytochrome b5 reductase and cytochrome b5. Quinones, such as the anticancer drug adriamycin (doxorubicin) and menadione, can undergo one-electron reduction catalyzed by NADPH cytochrome P-450 reductase. The semiquinone product may be oxidized back to the quinone with the concomitant production of superoxide anion radical, giving rise to redox cycling and potential cytotoxicity. This underlies the cardiac toxicity of adriamycin (see chap. 6). 

NADH cytochrome b5 reductase and cytochrome bs. The carbanion or carbene intermediates may rearrange with loss of a halogen ion (chap. 7, Fig. 77). 

The level of a particular enzyme involved in xenobiotic metabolism can obviously affect the extent of metabolism by that enzyme. Again, competition may play a part if endogenous and exogenous substrates are both metabolized by an enzyme, as is the case with some of the forms of cytochromes P-450, which metabolize steroids, or NADPH cytochrome P-450 reductase and cytochrome b5 reductase, which are also involved in heme catabolism and fatty acid metabolism, respectively. 

There are other components of the P-450 system, namely, NADPH cytochrome P-450 reductase, NADH cytochrome b5 reductase, and cytochrome b5. 

Desaturation requires the cooperative action of two enzymes Cytochrome b5 reductase and stearoyl-CoA desaturase, in addition to an electron carrier protein, cyto-... 

Enzyme complexes occur in the endoplasmic reticulum of animal cells that desaturate at A5 if there is a double bond at the A8 position, or at A6 if there is a double bond at the A9 position. These enzymes are different from each other and from the A9-desaturase discussed in the previous section, but the A5 and A6 desaturases do appear to utilize the same cytochrome b5 reductase and cytochrome b5 mentioned previously. Also present in the endoplasmic reticulum are enzymes that elongate saturated and unsaturated fatty acids by two carbons. As in the biosynthesis of palmitic acid, the fatty acid elongation system uses malonyl-CoA as a donor of the two-carbon unit. A combination of the desaturation and elongation enzymes allows for the biosynthesis of arachidonic acid and docosahexaenoic acid in the mammalian liver. As an example, the pathway by which linoleic acid is converted to arachidonic acid is shown in figure 18.17. Interestingly, cats are unable to synthesize arachidonic acid from linoleic acid. This may be why cats are carnivores and depend on other animals to make arachidonic acid for them. Also note that the elongation system in the endoplasmic reticulum is important for the conversion of palmitoyl-CoA to stearoyl-CoA. 

Hackett, C.S., Novoa, W.B., Kensil, C.R. Strittmatter, P. (1988). NADH binding to cytochrome b5 reductase blocks the acetylation of lysine 110. Journal of Biological Chemistry 263, 7539—43. 

Hackett, C.S. Strittmatter, P. (1984). Covalent cross-linking of the active sites of vesicle-bound cytochrome b5 and NADH-cytochrome b5 reductase. Journal of Biological Chemistry 259, 3275-82. 

Hyde, G.E. Campbell, W.H. (1990). High-level expression in Escherichia coli of the catalytically active flavin domain of corn leaf NADH-nitrate, reductase and its comparison to human NADH-cytochrome b5 reductase. Biochemical and Biophysical Research Communications 168, 1285-91. 

Shirabe, K., Yubisui, T., Nishino, T. Takeshita, M. (1991). Role of cysteine residues in human NADH-cytochrome b5 reductase studied by site-directed mutagenesis. Journal of Biological Chemistry 266, 7531-6. 

The next major step was the development of an assay to determine the functional identity of the deduced PDesat-TnAllZ desaturase. Initial efforts to develop an in vitro reconstitutive biochemical assay combined the recombinant PDesat-TnAllZ protein purified from E. coli with phospholipids and a biochemical fraction from beef liver containing cytochrome b5 reductase and cytochrome b5. These efforts were unsuccessful (Knipple and Roelofs, unpublished). 

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