Isolation, separation and quantification of metabolites of the carcinogenic polycyclic aromatic hydrocarbon dibenz(a,h)anthracene

Phosphorylation of rabbit liver cytochrome P-450 LM2 and its effect on monooxygenase activity

The phosphorylation of rabbit liver microsomal cytochrome P-450 LM2 by catalytic subunit of cyclic AMP-dependent protein kinase (W. Pyerin et al. (1983) Carcinogenesis 4, 573) has now been studied in detail with purified soluble form of cytochrome P-450 as well as with the purified protein incorporated into model membranes. The apparent Km values for P-450 of the phosphorylation reaction in all experimental systems were in a range of 2-8 microM, while the Vmax values were dependent on the state of P-450. Upon phosphorylation, the reconstituted enzyme activities with benzphetamine (N-demethylation) and 7-ethoxycoumarin (O-deethylation) as substrates were reduced to 30-40% of control.

Plasma membrane glycoproteins covalently bound to silica beads as a model for molecular studies of cell-cell interactions in culture.

Abstract In previous studies, we have shown that plasma membrane glycoproteins are of major importance in the density-dependent regulation of growth of normal diploid fibroblasts. Due to the hydrophobic portions of these molecules, functional studies in cell culture are often diffucult to perform and to interpret. Specially, the addition of these molecules in soluble form to cell culture, after depletion of detergents needed for their solubilization, leads to aggregation and internalization. Therefore, we developed a method for the covalent immobilization of the solubilized plasma membrane proteins to derivatized silica beads for further investigations on the molecular nature of the active …

Permissive and suppressive effects of dexamethasone on enzyme induction in hepatocyte co-cultures.

1. Steroids are known to act as permissive factors in hepatocytes. This study shows that dexamethasone (DEX) is a permissive factor for induction of CYP2B1/2, CYP3A1, CYP2A1 and probably also CYP2C11 in cultures with primary rat hepatocytes. 2. The induction factor of phenobarbital (PB)-induced formation of 16beta-hydroxytestosterone (OHT), a testosterone biotransformation product predominantly formed by CYP2B1, is increased 18-fold by the addition of 32 nM DEX to the culture medium. Interestingly, higher concentrations of DEX up to 1000 nM led to a concentration-dependent maximally 5-fold decrease (p = 0.002) of phenobarbital-induced 16beta-OHT formation compared with the effect observed w…

Mechanisms of Toxification and Detoxification which Challenge Drug Candidates and Drugs

Almost all drugs are metabolized in the human organism. In most cases this changes the toxicity, sometimes by toxification, sometimes by detoxification. For obvious ethical reasons, the toxicity cannot be experimentally studied in human beings. In systems available for toxicity studies such as whole animals or human or animal cells in culture, the drug metabolism is substantially different from that in the human organism. Risk assessment for human therefore requires knowledge of drug metabolism, its differences between systems, and the consequences for toxicity. In phase 1 of drug metabolism (oxidoreductions and hydrolyses) drugs are often toxified. This is especially the case if the result…

Detoxication and toxication of foreign compounds

Infiuence of Foreign Compounds on Formation and Disposition of Reactive Metabolites

Many toxic compounds are unreactive and need biotransformation in order to exert their toxic effects. Several enzymes control the formation or disposition of reactive metabolites. Especially well studied is the group of enzymes responsible for the control of reactive epoxides. Such epoxides may bind spontaneously to DNA, RNA and protein. These alterations of critical cellular macromolecules may disturb the normal biochemistry of the cell and lead to cytotoxic, allergenic, mutagenic and carcinogenic effects. Whether these effects will be manifested depends on the chemical reactivity as well as on other properties (geometry, lipophilicity) of the epoxide in question. Enzymes controlling the c…

Identification and Characterization of a Novel Epoxide Hydrolase From Mouse Liver Microsomes

A new microsomal epoxide hydrolase (mEH2) has been identified and characterized. This enzyme has properties which distinguish it from previously described cytosolic (cEH) or membrane-bound (mEH1) epoxide hydrolases. The enzyme is an integral microsomal protein which is not dissociated from the membrane by repeated washing, high ionic strength salt, or chaotropic agent solutions, or by sonication. It is very different from the normally described microsomal epoxide hydrolase (mEH1) as shown by its different substrate specificity and kinetic properties and by immunological criteria. In contrast to the hitherto described microsomal epoxide hydrolase, mEH1, the new enzyme effectively catalyzes t…