Resolution of mouse hepatic cytochrome P-450 isomers by chromatofocusing

Marriage, H.; Harvey, David J.

doi:10.1016/s0021-9673(01)87039-4

Cited by 12 publications

(2 citation statements)

References 25 publications

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“…The two small peaks between fractions I and II were contaminated with hemoglobin as determined spectrally, and the shoulder in the elution profile after fraction II represented about 7% of the applied P-450. The chromotofocusing procedure used is a modification of the system described by Marriage and Harvey (1986) for the resolution of mouse hepatic P-450 isozymes.…”

Section: Methodsmentioning

confidence: 99%

Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes

Ding¹,

Coon²

1988

Biochemistry

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Two forms of cytochrome P-450, designated P-450NMa and P-450NMb, were purified to electrophoretic homogeneity from rabbit nasal microsomes. The purified cytochromes, which contained 14-16 nmol of P-450/mg of protein, exhibited apparent monomeric molecular weights of 49,500 and 51,000, respectively. As indicated by several criteria, including the amino acid composition, absorption spectra, and peptide maps, the two nasal forms of P-450 are distinct from each other. Furthermore, as judged by the NH2-terminal amino acid sequences, they are distinct from all other P-450 cytochromes described to date. In the ferric form, P-450NMa is in the low-spin state, whereas P-450NMb is predominantly in the high-spin state. When reconstituted with NADPH-cytochrome P-450 reductase and phospholipid, P-450NMa is very active in the oxidation of ethanol as well as several nasal procarcinogens, including the N-deethylation of N-nitrosodiethylamine, the O-deethylation of phenacetin, and the N-demethylation of hexamethyl-phosphoramide. P-450NMb also metabolizes these substrates, but at lower rates. Both nasal forms are also active with testosterone, with P-450NMa oxidizing the substrate in the 17-position to give androstenedione and P-450NMb catalyzing hydroxylation in the 15 alpha-, 16 alpha-, and 19-positions. The two cytochromes represent the major portion of the total P-450 in nasal microsomes, but the corresponding forms could not be detected in hepatic microsomes.

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Section: Methodsmentioning

confidence: 99%

Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes

Ding¹,

Coon²

1988

Biochemistry

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“…These particular isoenzymes can be separated based on their isoelectric points. The application of chromatofocusing to separate isoenzymes has become increasingly popular (18,19). Proteins bind to this weak anionic exchanger at pH's that are higher than their isoelectric points.…”

Section: Resultsmentioning

confidence: 99%

Carboxylesterase isoenzyme specific deacylation of diacetoxyscirpenol (anguidine)

Wu¹,

Marletta²

1988

Chem. Res. Toxicol.

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The deacylation reaction of diacetoxyscirpenol (DAS), a trichothecene mycotoxin, was studied by using five carboxylesterase isoenzymes isolated from mouse liver microsomes. The simultaneous isolation of the five microsomal carboxylesterase isoenzymes was accomplished by chromatofocusing. DAS serves as a unique substrate since both acetyl groups are accessible for hydrolysis, providing an opportunity to investigate the regioselective hydrolysis of DAS with the various isoenzymes. A novel basic carboxylesterase isoenzyme, pI 8.4-8.2, was isolated. This isoenzyme exhibits strict regioselectivity toward DAS hydrolysis; only the acetyl group at C-4, but not that at C-15, is hydrolyzed. Moreover, this is the major isoenzyme responsible for the deacylation of DAS in mouse hepatic microsomes. The rate constant determined for deacylation of DAS at C-4 for this isoenzyme is 130-1000 times greater than that of the other carboxylesterases. This isoenzyme was also the most efficient for the hydrolysis of 4-monoacetoxyscirpenediol with a rate constant 30-100 times greater than that of the other isoenzymes.

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Stem cell‐derived cardiomyocytes and hepatocytes as tools for drug development and screening applications

Cameron

Marriage

Hay³

et al. 2015

Stem Cells in Regenerative Medicine

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Resolution of mouse hepatic cytochrome P-450 isomers by chromatofocusing

Cited by 12 publications

References 25 publications

Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes

Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes

Carboxylesterase isoenzyme specific deacylation of diacetoxyscirpenol (anguidine)

Stem cell‐derived cardiomyocytes and hepatocytes as tools for drug development and screening applications

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