Membranes metabolic enzyme systems

Orrenius, S., J.L. Ericsson, and L. Ernster (1965). Phenobarbital-induced synthesis of the microsomal drug-metabolizing enzyme system and its relationship to the proliferation of endoplasmic membranes. A morphological and biochemical study. J. Cell. Biol. 25, 627-639. 

The endothelial cells actively, as well as passively, serve to protect the brain. Because they contain a variety of drug-metabolizing enzyme systems similar to the drug-metabolizing enzymes found in the liver, the endothelial cells can metabolize neurotransmitters and toxic chemicals and, therefore, form an enzymatic barrier to entry of these potentially harmful substances into the brain. They actively pump hydrophobic molecules that diffuse into endothelial cells back into the blood (especially xenobiotics) with P-glycoproteins, which act as transmembranous, ATP-dependent efflux pumps. Although lipophilic substances, water, oxygen, and carbon dioxide can readily cross the blood-brain barrier by passive diffusion, other molecules depend on specific transport systems. Differential transporters on the luminal and abluminal endothelial membranes can transport compounds into, as well as out of, the brain. 

FIGURE 18.5 Schematic representation of types of multienzyme systems carrying out a metabolic pathway (a) Physically separate, soluble enzymes with diffusing intermediates, (b) A multienzyme complex. Substrate enters the complex, becomes covalently bound and then sequentially modified by enzymes Ei to E5 before product is released. No intermediates are free to diffuse away, (c) A membrane-bound multienzyme system. 

The water-soluble vitamins generally function as cofactors for metabolism enzymes such as those involved in the production of energy from carbohydrates and fats. Their members consist of vitamin C and vitamin B complex which include thiamine, riboflavin (vitamin B2), nicotinic acid, pyridoxine, pantothenic acid, folic acid, cobalamin (vitamin B12), inositol, and biotin. A number of recent publications have demonstrated that vitamin carriers can transport various types of water-soluble vitamins, but the carrier-mediated systems seem negligible for the membrane transport of fat-soluble vitamins such as vitamin A, D, E, and K. 

During the last ten years, it has become apparent that calcium-dependent papain-like peptidases called calpains (EC 3.4.22.17) represent an important intracellular nonlysosomal enzyme system [35][36], These enzymes show limited proteolytic activity at neutral pH and are present in virtually every eukaryotic cell type. They have been found to function in specific proteolytic events that alter intracellular metabolism and structure, rather than in general turnover of intracellular proteins. Calpains are composed of two nonidentical subunits, each of which contains functional calcium-binding sites. Two types of calpains, i.e., /i-calpain and m-calpain (formerly calpain I and calpain II, respectively), have been identified that differ in their Ca2+ requirement for activation. The activity of calpains is regulated by intracellular Ca2+ levels. At elevated cytoplasmic calcium concentrations, the precursor procal-pain associates with the inner surface of the cell membrane. This interaction seems to trigger autoproteolysis of procalpain, and active calpain is released into the cytoplasm [37]. 

The endoplasmic reticulum (ER) is an extensive closed membrane system consisting of tubular and saccular structures. In the area of the nucleus, the ER turns into the external nuclear membrane. Morphologically, a distinction is made between the rough ER (rER) and the smooth ER (sER). Large numbers of ribosomes are found on the membranes of the rER, which are lacking on the sER. On the other hand, the sER is rich in membrane-bound enzymes, which catalyze partial reactions in the lipid metabolism as well as biotransformations. 

The cytochrome P450 system is the principal enzyme system for the metabolism of lipophilic xenobiotics. It is a heme-containing, membrane-bound, multi-enzyme system which is present in many tissues in vivo but is present at the highest level in liver. A coenzyme, cytochrome P450 NADPH oxidoreductase (OR), is essential for P450 catalytic function and cytochrome bs may stimulate catalytic activities of some enzymes. In human liver, it is estimated that there are 15-20 different xenobiotic-metabolizing cytochrome P450 forms. A standard nomenclature, based on relatedness of the amino acid sequences, has been developed (Nelson et al., 1993). The most recent... 

Tissue lysate (or homogenates), post-mitochondrial supernatants and microsomes offer several practical advantages for the study of xenobiotic metabolism. The principal advantages are that the human tissues provide a complete system containing all the enzymes in ratios found in vivo, and tissue fractions are stable in relatively long-term storage. Within the different types of tissue fractions, microsomes provide an enrichment of the membrane-bound enzymes, and post-mitochondrial supernatants provide a means to study both membrane-bound and soluble enzymes. Tissue fractions are easily prepared from a variety of tissues including human liver and can be cryopreserved for several years. This allows detailed characterization of the tissue prior to use with xenobiotics of unknown routes of metabolism... 

In the induction of enzymes of galactose metabolism in E. coli, three enzymes are involved -galactosidase (which catalyses the hydrolysis of the y -glycosidic bonds of lactose), galactose permease (which is responsible for transport of lactose across the cell membrane) and a third enzyme, A-protein, apparently not directly involved in galactose metabolism. The system has an environmental inducer, galactose, and in its presence the number of /)-galactosidase molecules rises from 5-10 to 10,000 within the cell. The addition of the inducer can increase the protein production in less than five minutes after its addition. Protein synthesis of these enzymes stops almost immediately in the absence of lactose. 

Kinetic toxicology, also known as metabolic or pharmacologic toxicology, involves toxicants that are transported and metabolized in the body. Such substances are called systemic poisons and they are studied under the discipline of systemic toxicology. Systemic poisons may cross cell membranes (see Chapter 3) and act on receptors such as cell membranes, bodies in the cells, and specific enzyme systems. The effect is dose responsive, and it is terminated by processes that may include metabolic conversion of the toxicant to a metabolic product, chemical binding, storage, and excretion from the organism. 

Studies with various subcellular fractions are useful to ascertain which enzyme systems are involved in the metabolism of a chug candidate. In the absence of added cofactors, oxidative reactions such as oxidative deamination that are supported by mitochondria or by Ever microsomes contaminated with mitochondria membranes (as is the case with microsomes prepared from frozen liver samples) are likely catalyzed by monoamine oxidase (MAO), whereas oxidative reactions supported by cytosol are likely catalyzed by aldehyde oxidase and/or xanthine oxidase (a possible role for these enzymes in the metabolism of... 

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