Alkaline Density Gradient Floatation of Membranes: Polypeptide Composition of the Mammalian Peroxisomal Membrane
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- 作者：Gouveia ; Alexandra M. M. ; Reguenga ; Carlos ; Oliveira ; Má ; rcia E. M. ; Eckerskorn ; Christoph ; et. al.
- 刊名：Analytical Biochemistry
- 出版年：1999
- 出版时间：October 15, 1999
- 年：1999
- 卷：274
- 期：2
- 页码：270-277
- 全文大小：202 K
文摘
A method for purification of the peroxisomal membrane from rat liver is described. The procedure consists of floating the (contaminated) peroxisomal membranes through an alkaline sucrose density gradient. A good resolution between the peroxisomal membrane and other membrane systems is achieved. Using these floated peroxisomal membranes we have determined that only 7.8 ± 0.9 % of the total peroxisomal protein is alkali resistant. The polypeptide composition of these highly pure peroxisomal membranes was analyzed by SDS–PAGE. Bands corresponding to polypeptides with apparent molecular masses of 15, 18, 22, 24, 26, 29, 35, 36, 38, 40, 52, 55, 70, 74–77, and 88 kDa are detected upon Coomassie blue staining of polyacrylamide gels. The identity of several of these polypeptides was determined by N-terminal sequencing and Western blotting analysis.