Preparation and Purification of Peroxisomes; Subfractionation of Purified Peroxisomes; Acyl-CoA Oxidase Activity Measurements
The isolation of peroxisomes is generally accomplished in three steps. After homogenisation of the tissue or disruption of the cells, a subcellular fraction enriched in peroxisomes is obtained by differential centrifugation (for a comprehensive treatise on centrifugation, see ). This fraction is then further separated by means of density gradient centrifugation. Such a procedure will be demonstrated on liver, obtained from control rats and from rats treated with bezafibrate. Hypolipidemic drugs such as bezafibrate cause a pronounced proliferation of peroxisomes in liver and kidney of rodents. This is associated with hepatomegaly and the induction of a particular set of peroxisomal enzymes involved in the β-oxidation of fatty acids  (see Exp. 1.3).
KeywordsPeroxisomal Membrane Urate Oxidase Peroxisomal Fraction Bile Acid Intermediate Nycodenz Gradient
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- 2.De Duve C, Pressman BC, Gianetto R, Wattiaux R, Appelmans F (1955) Tissue fractionation studies. 6. Intracellular distribution patterns of enzymes in rat liver tissue. Biochem J 60:604–617Google Scholar
- 3.Evans WH (1982) Subcellular membranes and isolated organelles: preparative techniques and criteria for purity. Techniques in Lipid and Membrane Biochemistry B407a: 1–46Google Scholar
- 10.Rickwood D (1989) Centrifugation — A practical approach (2nd Ed.) Practical Approach Series. IRL Press, Oxford.Google Scholar
- 11.Van Veldhoven PP, Mannaerts GP (1993) Assembly of peroxisomal membranes. In: Eds. Maddy AH and Harris JR. Subcellular Biochemistry, membrane biogenesis. Plenum Publishing Press, New York (in press)Google Scholar
- 3.Kamijo K, Taketani S, Yokota S, Osumi T, Hashimoto T (1990) The 70-kDa peroxisomal membrane protein is a member of the Mdr (P-glycoprotein)-related ATP binding protein superfamily. J Biol Chem 265:4531–4540Google Scholar
- 5.Schepers L, Van Veldhoven PP, Casteels M, Eysen HJ, Mannaerts GP (1990). Presence of three acyl-CoA oxidases in rat liver peroxisomes. An inducible fatty acyl-CoA oxidase, a non-inducible fatty acyl-CoA oxidase and a non-inducible trihydroxycoprostanoyl-CoA oxidase. J Biol Chem 265:5242 – 5246PubMedGoogle Scholar
- 8.Van Veldhoven PP, Vanhove G, Asselberghs S, Eyssen HJ, Mannaerts GP (1992) Substrate specificities of rat liver peroxisomal acyl-CoA oxidases: Palmitoyl-CoA oxidase (inducible acyl-CoA oxidase), pristanoyl-CoA oxidase (non-inducible acyl-CoA oxidase) and trihydroxycoprostanoyl-CoA oxidase. J Biol Chem 267:20065 – 20074PubMedGoogle Scholar