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Silke Gehrhardt, Eva Blume, Gerhard A. Cumme, Renate Bublitz, Heidrun Rhode, Anton Horn
Gel Chromatographic Characterization of the Hydrophobic Interaction of Glycosylphosphatidylinositol-Alkaline Phosphatase with Detergents
The interaction of a glycosylphosphatidylinositol (GPI) protein with different detergents was studied for the first time with a purified protein. Four differently hydrophobic fractions of GPIalkaline phosphatase (GPIAP) from calf intestine were used as model proteins. The mode of interaction was determined by investigating (i) the selfaggregation behaviour of the GPIAP fractions, (ii) the interference of detergents with GPIAP binding to octylSepharose, and (iii) the elution of GPIAP bound to octylSepharose. It was shown that polyoxyethylenetype detergents surprisingly interact much stronger than noctylglucoside with GPIAP, which is in contrast to the known behaviour of GPIproteins in natural membranes. Gel filtration chromatography of Triton X-100 at concentrations above the critical micellar concentration yields three different micelle species with apparent molecular weights of about 166, 54, and 16 kDa. GPIAP fraction II, which is shown to bear only one anchor per dimer, does not bind to any of these micelles. We demonstrate that a complex is formed containing about 150 Triton X-100 molecules and about 4700 molecules of water per molecule of GPIAP dimer. The experimental findings are in accordance with a simple geometrical model based on the physical data of fatty acids and the arrangement, mean size, and shape of Triton X-100 molecules.
Biological Chemistry, Walter de Gruyter
Print ISSN: 1431-6730
Volume: 381, 02/2000
Pages: 161 - 172