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The occurrence of multiple forms of cytochrome P-450 in hepatic microsomes from untreated rats and mice
Abstract:
The hepatic microsomes of rat and mice were subfractionated by the procedure of Dallner. When a 1.3 M sucrose lower layer was used for the two-step discontinuous gradient, no differences in spectral characteristics were noted between subfractions, though the smooth fractions (SER) had higher oxidative activity towards the substrates tested. When lower layers of 1.05, 1.1 or 1.15 M sucrose were used, and the SER isolated contained cytochdrome P-450 with significantly different spectral characteristics from that of the rough fraction (RER). The SER cytochrome P-450 had a wavelength maximum in the carbon-monoxide reduced difference spectrum that was significantly lower (ca. 1.0 nm) than that in the RER. In addition, the type I:CO-reduced spectral ratio of these fractions is significantly elevated. These data indicate that liver microsomes from untreated rats and mice contain more than one cytochrome P-450 and that of these cytochromes may be located in different parts of the endoplasmic reticulum.
Insights
Liver microsomes contain multiple cytochrome P-450 forms. These distinct cytochrome P-450 variants are differentially distributed within the smooth and rough endoplasmic reticulum fractions.
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Hepatic microsomes are crucial for drug metabolism.
- Cytochrome P-450 enzymes within microsomes exhibit diverse functions.
- Understanding cytochrome P-450 localization is key to drug metabolism studies.
Purpose of the Study:
- To investigate the heterogeneity of cytochrome P-450 in rat and mouse liver microsomes.
- To determine if different cytochrome P-450 forms are localized in smooth (SER) and rough (RER) endoplasmic reticulum.
Main Methods:
- Subfractionation of hepatic microsomes using Dallner's procedure.
- Two-step discontinuous sucrose gradient centrifugation.
- Spectral analysis of cytochrome P-450 in isolated SER and RER fractions.
Main Results:
- No spectral differences in cytochrome P-450 were observed between SER and RER when using a 1.3 M sucrose layer.
- Using lower sucrose concentrations (1.05-1.15 M) revealed distinct spectral characteristics for SER cytochrome P-450 compared to RER.
- SER cytochrome P-450 showed a lower wavelength maximum and elevated Type I:CO-reduced spectral ratio.
Conclusions:
- Liver microsomes contain multiple forms of cytochrome P-450.
- These cytochrome P-450 forms exhibit differential localization within the endoplasmic reticulum.
- The findings suggest distinct functional roles for cytochrome P-450 in SER and RER.