Cytochrome defect

The well-known fact that in irreversibly damaged cells, respiratory control is lost and is accompanied by oxidation of cytochromes a and as, as well as NADH (Taegtmeyer et al., 1985), was originally thoug it to be due to substrate deficiency (Chance and Williams, 1955) but may be due to an enzymatic defect resulting in an inability to metabolize NADH-linked substrates (Pelican etal., 1987). It seems likely therefore that return of function is dependent on preservation of mitochondrial membrane integrity, and the structure and activities of respiratory chain (R.C) complexes I-IV (Chance and Williams, 1955). 

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). 

Abnormalities of the respiratoiy chain. These are increasingly identified as the hallmark of mitochondrial diseases or mitochondrial encephalomyopathies [13]. They can be identified on the basis of polarographic studies showing differential impairment in the ability of isolated intact mitochondria to use different substrates. For example, defective respiration with NAD-dependent substrates, such as pyruvate and malate, but normal respiration with FAD-dependent substrates, such as succinate, suggests an isolated defect of complex I (Fig. 42-3). However, defective respiration with both types of substrates in the presence of normal cytochrome c oxidase activity, also termed complex IV, localizes the lesions to complex III (Fig. 42-3). Because frozen muscle is much more commonly available than fresh tissue, electron transport is usually measured through discrete portions of the respiratory chain. Thus, isolated defects of NADH-cytochrome c reductase, or NADH-coenzyme Q (CoQ) reductase suggest a problem within complex I, while a simultaneous defect of NADH and succinate-cytochrome c reductase activities points to a biochemical error in complex III (Fig. 42-3). Isolated defects of complex III can be confirmed by measuring reduced CoQ-cytochrome c reductase activity. 

Defects of complex II. These have not been fully characterized in the few reported patients, and the diagnosis has often been based solely on a decrease of succinate-cytochrome c reductase activity (Fig. 42-3). However, partial complex II deficiency was documented in muscle and cultured fibroblasts from two sisters with clinical and neuroradiological evidence of Leigh s syndrome, and molecular genetic analysis showed that both patients were homozygous for a point mutation in the flavoprotein subunit of the complex [17]. This was the first documentation of a molecular defect in the nuclear genome associated with a respiratory chain disorder. 

Extensive studies into the association of this cytochrome b with CGD neutrophils were performed by Segal and Jones, and by other workers, in the late 1970s and early 1980s. The cytochrome was completely absent (as determined by the absence of a distinctive absorption spectrum in spectroscopic studies) in almost all cases of X-linked CGD, but present at decreased levels in female relatives of these patients. In almost all cases of autosomal recessive CGD, the cytochrome was present but non-functional, in that it did not become reduced upon cellular activation. This indicated both the heterogeneous nature of the disease and also that some other biochemical defect was responsible for impaired function in these patients. Hence, the search was on for other components of the NADPH oxidase. 

Thus, it has been shown that, in the majority of X-linked CGD patients, the abnormality is due to the failure to transcribe the mRNA encoding the large (/J) subunit of the b cytochrome. In these patients, both the heavy /)-chain and the light a-chain are absent, even though the molecular defect appears to be restricted to the heavy chain thus, expression of the heavy chain is somehow necessary for the expression/translation/stabilisation of the... 

The second major breakthrough in understanding the defect in CGD neutrophils came through the development of assays in which the NADPH oxidase can be activated in a cell-free system in vitro ( 5.3.2.3). In these systems, activation of the oxidase can be achieved by the addition of cytoplasm to plasma membranes in the presence of NADPH and arachidonic acid (or SDS or related substances). Interestingly, the oxidase cannot be activated in these cell-free systems using extracts from CGD neutrophils however, cytosol and plasma membranes from normal and CGD neutrophils may be mixed, and in most cases activity is restored if the correct mixing pattern is used. For example, as may be predicted, in X-linked CGD it is the membranes that are defective (because the cytochrome b is deficient), whereas in autosomal recessive CGD the cytosol is defective in the cell-free system. 

Cumutte, J. T., Scott, P. J., Babior, B. M. (1989). Functional defect in neutrophil cytosols from two patients with autosomal recessive cytochrome-positive chronic granulomatous disease. J. Clin. Invest. 83, 1236-40. 

Segal, A. W. (1988). Cytochrome b.245 and its involvement in the molecular pathology of chronic granulomatous disease. In Phagocytic Defects, vol. II (Hematology/Oncology Clinics of North America), pp. 213-23, W. B. Saunders, Philadelphia. 

Haem synthesis is a good example of a pathway that is partly compartmentalized. The pathway (Figure 5.16) occurs in all cell types for the production of respiratory cytochromes and begins within mitochondria but the majority of the reactions occur in the cytosol cell. Because mature red cells have no subcellular organelles, haem synthesis occurs only in early RBC progenitor cells. Although this is a relatively simple pathway, there are a number of well-known enzyme defects that cause a group of diseases called the porphyrias. 

Caspase-8 activates the effector caspases either directly, or indirectly by promoting the cytochrome c (see p. 140) from mitochondria. Once in the cytoplasm, cytochrome c binds to and activates the protein Apaf-1 (not shown) and thus triggers the caspase cascade. Apoptotic signals can also come from the cell nucleus. If irreparable DNA damage is present, the p53 protein (see p. 394)—the product of a tumor suppressor gene—promotes apoptosis and thus helps eliminate the defective cell. 

Cytochrome P450 inducer (oral contraceptive failure) Autoinduction Rare blood cell dyscrasias aplastic anemia, agranulocytosis Hepatotoxicity Rash risk, including Stevens-Johnson syndrome Risk for SIADH Teratogenicity risk neural tube defects, craniofacial defects... 

Zumft, W. G., Dohler, K., Ktirner, H., Lochelt, S., Viebrock, A., and Frunzke, K. (1988). Defects in cytochrome cd, dependent nitrite respiration of transposon Tn5-induced mutants from Pseudomonas stutzeri. Arch. Microbiol. 149, 492-498. 

NADH coenzyme Q reductase defect (complex I) Succinate coenzyme Q reductase defect (complex II) Coenzyme Q cytochrome C reductase defect (complex III)... 

This is a sedative drug with low adult toxicity, which proved to be a very potent human teratogen, causing phocomelia (shortening of the limbs) and other defects when taken between the third and eighth week. In some cases, only a few doses were taken, but on the critical days (e.g., days 24-27 for phocomelia of arms). It is not readily reproducible in laboratory animals (e.g., rats). Mechanism is unknown, but a metabolite suspected, possibly produced by cytochrome P-450. A number of metabolites are produced and some chemical breakdown occurs. Phthalylglutamic acid metabolite is teratogenic in mice. Thalidomide may acylate nucleic acids and polyamines. The S-enantiomer is more embryotoxic than the R-enantiomer. 

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