scholarly journals Cytochrome P-450 induction by clofibrate. Purification and properties of a hepatic cytochrome P-450 relatively specific for the 12- and 11-hydroxylation of dodecanoic acid (lauric acid)

1982 ◽  
Vol 203 (1) ◽  
pp. 161-168 ◽  
Author(s):  
G. Gordon Gibson ◽  
Terry C. Orton ◽  
Paul P. Tamburini
Keyword(s):  
Endoplasmic Reticulum ◽  
Lauric Acid ◽  
Dodecanoic Acid ◽  
Specific Content ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Mixed Function Oxidase System ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

Hypolipidaemic drugs induce peroxisomal proliferation in the liver and many induce the formation of the hepatic endoplasmic reticulum in general and the formation of cytochrome P-450 in particular. We have induced the formation of rat liver microsomal cytochrome P-450 by the administration of the hypolipidaemic drug clofibrate, isolated the endoplasmic reticulum, solubilized the cytochrome P-450 from these membranes and subdivided the cytochrome P-450 into four fractions by the use of hydrophobic, anionic, cationic and adsorption chromatography. One of these fractions (cytochrome P-450 fraction 1) was highly purified to a specific content of 17nmol of cytochrome P-450/mg of protein and the protein was active in a reconstituted enzyme system towards the 12- and 11-hydroxylation of the fatty acid, dodecanoic (lauric) acid, with preferential activity towards the 12-hydroxy metabolite. This reconstituted activity was absolutely dependent on NADPH, NADPH-cytochrome P-450 reductase and cytochrome P-450, indicating the role of the mixed-function oxidase system in the metabolism of lauric acid. Another fraction of the haemoprotein (cytochrome P-450 fraction 2) preferentially formed 11-hydroxylauric acid, whereas a third fraction (cytochrome P-450 fraction 3) exhibited only trace laurate oxidase activity and was similar to the phenobarbitone form of the haemoprotein in that these last two cytochromes rapidly turned-over the drug benzphetamine. The molecular weights and spectral properties of these cytochrome P-450 fractions are reported, along with the phenobarbitone-induced form of the enzyme and the nature of the cytochrome(s) induced by clofibrate pretreatment are discussed in the terms of possible haemoprotein heterogeneity.

scholarly journals Decrease in hepatic cytochrome P-450 by cobalt. Evidence for a role of cobalt protoporphyrin

1982 ◽  
Vol 204 (1) ◽  
pp. 103-109 ◽  
Author(s):  
J F Sinclair ◽  
P R Sinclair ◽  
J F Healey ◽  
E L Smith ◽  
H L Bonkowsky
Keyword(s):  
Chick Embryo ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Cobalt Protoporphyrin ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

Exposure of cultured chick-embryo hepatocytes to increasing concentrations of CoCl2 in the presence of allylisopropylacetamide results in formation of cobalt protoporphyrin, with a reciprocal decrease in haem and cytochrome P-450. Treatment of rats with CoCl2 (84 mumol/kg) and 5-aminolaevulinate (0.2 mmol/kg) also results in formation of cobalt protoporphyrin and a decrease in cytochrome P-450 in the liver. Hepatic microsomal fractions from rats treated with phenobarbital, CoCl2 and 5-aminolaevulinate were analysed by polyacrylamide gel electrophoresis. Cobalt protoporphyrin was associated mainly with proteins of 50000-53000 mol.wt. The results suggest that the formation of cobalt protoporphyrin occurred at the expense of the synthesis of haem, leading to a decrease in cytochrome P-450. Furthermore, the cobalt protoporphyrin that was formed may itself have been incorporated into apocytochrome P-450.

Regulation of hepatic cytochrome P-450 during Infectious disease

1990 ◽  
Vol 68 (6) ◽  
pp. 777-781 ◽  
Author(s):  
Kenneth W. Renton ◽  
Leah C. Knickle
Keyword(s):  
Infectious Disease ◽  
Viral Infections ◽  
Oligonucleotide Probe ◽  
Protein S ◽  
Unique Sequence ◽  
Mixed Function Oxidase ◽  
Oxidase System ◽  
Mixed Function Oxidase System ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

During episodes of infectious disease the mixed function oxidase system is depressed and the capacity of the liver to metabolize drags can be compromised in both animals and humans. The depression that occurs during viral infections is mediated via the production of interferon. This action of interferon requires the synthesis of an intermediate protein(s) yet to be identified. Using an oligonucleotide probe for a unique sequence in cytochrome P-450LAω we have now shown that the mRNA for this isozyme is depressed following the administration of interferon inducers. The magnitude in the loss of mRNA corresponds to the magnitude of the loss in the levels of this isozyme. This depression is observed within 6 h of interferon exposure. It is concluded that the decrease in drag metabolism during viral infections is caused by an interferon-mediated loss in mRNA and subsequent cytochrome P-450 synthesis in the liver.Key words: cytochrome P-450, drug metabolism, mixed function oxidase, interferon, viral infection.

The Role of Membrane Proteins and Phospholipids in the Interaction of Ribosomes with Endoplasmic Reticulum Membranes

1975 ◽  
Vol 53 (9) ◽  
pp. 1039-1045 ◽  
Author(s):  
Serge Jothy ◽  
Jean-Louis Bilodeau ◽  
Henry Simpkins
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Rat Liver ◽  
Rough Endoplasmic Reticulum ◽  
Binding Sites ◽  
Molecular Weights ◽  
Low Concentrations ◽  
Phospholipid Headgroups ◽  
Hydrolysis Of

Hydrolysis of the membrane proteins and phospholipid headgroups of rat liver rough endoplasmic reticulum membranes showed that the ribosomal binding sites involve membrane proteins susceptible to low concentrations of trypsin, chymotrypsin, and papain. Three membrane proteins having molecular weights of 120 000, 93 000 and 36 000 are found to be altered by trypsin and chymotrypsin treatment. Also the polar headgroup of phosphatidylinositol appears to play a role in the binding process.

scholarly journals Role of haem in the synthesis and assembly of cytochrome P-450

1977 ◽  
Vol 164 (2) ◽  
pp. 295-303 ◽  
Author(s):  
Kolari S. Bhat ◽  
Mohinder K. Sardana ◽  
Govindarajan Padmanaban
Keyword(s):  
Endoplasmic Reticulum ◽  
Binding Capacity ◽  
Direct Interaction ◽  
Normal Animal ◽  
Functional Protein ◽  
A Site ◽  
Rate Limiting ◽  
Cytochrome P 450

By using 3-amino-1,2,4-triazole, an inhibitor of haem synthesis, and 2-allyl-2-isopropylacetamide, a drug that degrades the haem moiety of cytochrome P-450, the involvement of haem in cytochrome P-450 synthesis and assembly was investigated. Phenobarbital was used to stimulate apo-(cytochrome P-450) synthesis. Degradation of preformed cytochrome P-450 haem does not result in a concomitant release of the apoprotein from the endoplasmic reticulum. The availability of haem for cytochrome P-450 synthesis in the normal animal is not rate-limiting. Prolonged inhibition of haem synthesis in vivo decreases the rate of apo-(cytochrome P-450) synthesis, although this effect is not discernible under conditions of short-term inhibition of haem synthesis. Under the former conditions exogenous haemin is able to counteract the decrease in the rate of apoprotein synthesis. In animals receiving successive injections of phenobarbital plus 3-amino-1,2,4-triazole, compared with those receiving phenobarbital only, the holo-(cytochrome P-450) content measured spectrally shows a greater decrease than could be accounted for by the decrease in the content of the total apoprotein. In addition to less haem being available under these conditions, the free apoprotein appears to have undergone some modification, such that its haem-binding capacity is considerably decreased. This particular effect could be due to a direct interaction of 3-amino-1,2,4-triazole or its metabolites with cytochrome P-450 rather than a consequence of haem deficiency. Apo-(cytochrome P-450) is capable of binding to the endoplasmic reticulum in a form and at a site, which can be reconstituted with haemin to yield the functional protein.

scholarly journals The role of 5-aminolaevulinate synthase, haem oxygenase and ligand formation in the mechanism of maintenance of cytochrome P-450 concentration in hepatocyte culture

1983 ◽  
Vol 210 (3) ◽  
pp. 855-857 ◽  
Author(s):  
L J Hockin ◽  
A J Paine
Keyword(s):  
Hepatocyte Culture ◽  
Cultured Hepatocytes ◽  
Pyridine Ligands ◽  
Haem Oxygenase ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

The present work shows that the ability of pyridines e.g. metyrapone, to maintain the cytochrome P-450 concentration in cultured hepatocytes is not due to their ability to alter the 5-aminolaevulinate synthase and haem oxygenase activities of the hepatocytes. Since ligands such as metyrapone will prevent the cobalt-mediated loss of hepatic cytochrome P-450 in rats, the hypothesis that ligand formation is the mechanism of maintenance of the cytochrome in hepatocyte culture was tested. The observation that non-pyridine ligands will maintain the cytochrome P-450 concentration supports this hypothesis.

Inhibition of the synthesis of hepatic cytochrome P-450 by the interferon-inducing agent poly rI.rC

1984 ◽  
Vol 62 (4) ◽  
pp. 379-383 ◽  
Author(s):  
Gurmit Singh ◽  
Kenneth W. Renton
Keyword(s):  
Amino Acids ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Microsomal Protein ◽  
Interferon Inducer ◽  
Molecular Weights ◽  
Drug Biotransformation ◽  
Mixed Function Oxidase System ◽  
Pyrene Hydroxylase ◽  
Cytochrome P 450

The levels of cytochrome P-450, cytochrome b5, aminopyrine N-demethylase, and benzo[a]pyrene hydroxylase were depressed in hepatic microsomes following treatment of mice with the interferon inducer poly rI.rC. The decrease in the hepatic mixed function oxidase system was accompanied by an increase in the incorporation of amino acids into total microsomal protein. Fractionation of solubilized microsomes using a Sephacryl S-200 gel filtration column demonstrated that the increase in amino acid incorporation tended to be associated with proteins with molecular weights under 67 000. The fractions which contained cytochrome P-450 were further separated using a DEAE cellulose column. The amount of labelled amino acids associated with the cytochrome P-450 fractions was uniformly depressed in preparations from poly rI.rC treated animals compared with saline-treated controls. These results suggest that poly rI.rC causes a depression in the rate of synthesis of the apoprotein of cytochrome P-450 while increasing the incorporation of amino acids into other hepatic proteins. The decrease in apocytochrome P-450 synthesis explains the marked loss of drug biotransformation which occurs following induction of interferon.

scholarly journals Role of haem in the induction of cytochrome P-450 by phenobarbitone. Studies in chick embryos in ovo and in cultured chick embryo hepatocytes

1981 ◽  
Vol 198 (2) ◽  
pp. 321-329 ◽  
Author(s):  
U Giger ◽  
U A Meyer
Keyword(s):  
Chick Embryo ◽  
Inhibitory Effect ◽  
Chick Embryos ◽  
In Ovo ◽  
Haem Biosynthesis ◽  
Rate Limiting ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

The role of haem synthesis during induction of hepatic cytochrome P-450 haemoproteins was studied in chick embryo in ovo and in chick embryos hepatocytes cultured under chemically defined conditions. 1. Phenobarbitone caused a prompt increase in the activity of 5-aminolaevulinate synthase, the rate-limiting enzyme of haem biosynthesis, and in the concentration of cytochrome P-450. This induction response occurred without measurable initial destruction of the haem moiety of cytochrome P-450. 2. When intracellular haem availability was enhanced by exogenous haem or 5-aminolaevulinate, phenobarbitone-medicated induction of cytochrome P-450 was not affected in spite of the well known repression of 5-aminolaevulinate synthase by haem. These data are consistent with the concept that haem does not regulate the synthesis of cytochrome P-450 haemoproteins. 3. Acetate inhibited haem biosynthesis at the level of 5-aminolaevulinate formation. When intracellular haem availability was diminished by treatment with acetate, phenobarbitone-medicated induction was decreased. 4. This inhibitory effect of acetate on cytochrome P-450 induction was reversed by exogenous haem or its precursor 5-aminolaevulinate. These data suggest that inhibition of haem biosynthesis does not decrease synthesis of apo-cytochrome P-450. Moreover, they indicate that exogenous haem can be incorporated into newly formed aop-cytochrome P-450.

scholarly journals Regulation of phenobarbital-inducible cytochrome P-450s in rat and mouse liver following dexamethasone administration and hypophysectomy

1988 ◽  
Vol 254 (3) ◽  
pp. 789-797 ◽  
Author(s):  
R R Meehan ◽  
L M Forrester ◽  
K Stevenson ◽  
N D Hastie ◽  
A Buchmann ◽  
...  
Keyword(s):  
Species Difference ◽  
Potent Inducer ◽  
The Difference ◽  
The Stability ◽  
And Function ◽  
Synthetic Glucocorticoids ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

Cytochrome P-450s are a superfamily of haem-containing proteins involved in the metabolism of foreign compounds, as well as a variety of endogenous molecules. The hepatic levels and function of this diverse group of enzymes are determined by both constitutive and xenobiotic regulators. To examine the role of constitutive factors in cytochrome P-450 regulation, the levels of three distinct groups of phenobarbital-inducible hepatic cytochrome P-450s were studied following dexamethasone-treatment or hypophysectomy. In the mouse, dexamethasone was a potent inducer of proteins within the PB1 (subfamily IIC), PB2c (family III) and PB3 (subfamily IIB) families. These findings were strikingly different from the effects in the rat where essentially no effect on PB3 expression and indeed suppression of proteins related to PB1 was observed. Determination of mRNA concentration indicated that the difference was at the level of transcription. These findings indicate that synthetic glucocorticoids have the potential to be potent phenobarbital-like inducing agents. In the mouse hypophysectomy, like dexamethasone, induced hepatic mRNA of P-450 from families P-450IIB, P-450IIC and P-450III. Again a species difference was observed as this treatment had essentially no effect in the rat. These data in the mouse indicate that factors produced in the pituitary can either affect the transcription rate of phenobarbital and dexamethasone-inducible P-450 genes or influence the stability of their mRNAs.

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