Amm. Gouveia et al., Alkaline density gradient floatation of membranes: Polypeptide compositionof the mammalian peroxisomal membrane, ANALYT BIOC, 274(2), 1999, pp. 270-277
A method for purification of the peroxisomal membrane from rat liver is des
cribed. The procedure consists of floating the (contaminated) peroxisomal m
embranes through an alkaline sucrose density gradient. A good resolution be
tween the peroxisomal membrane and other membrane systems is achieved. Usin
g 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 an
d Western blotting analysis. (C) 1999 Academic Press.