Different methods of membrane domains isolation result in similar 2-D distribution patterns of membrane domain proteins.
Membrane domains are highly specialized parts of the cell plasma membrane, carrying on and augmenting the incoming signals. To study their structural and functional properties, it is crucial to find the least damaging mode of their isolation. Using two different cell lines, epithelial HEK cells (clone E2M11) and S49 lymphoma cells, three methods of membrane domain isolation (i.e., detergent extraction, alkaline treatment, and "drastic" homogenization) were tested for similarity and reproducibility by 2-D electrophoresis. Our data show that the protein composition of membrane domains obtained by different isolation methods is similar and that approximately 60% of the spots are present in all membrane domain preparations. Furthermore, the same degree of similarity of 2-D profiles of the most intensively silver stained spots found in membrane domains of the two cell lines derived from different tissues suggests that the composition of a large part of membrane domains proteins is conservative. We suggest that these proteins may either be involved in the organization of membrane domain structure or represent the conservative component of signal transduction machinery.
Matousek
Petr
P
Institute fo Physiology, Academy of Sciences of the Czech Republic, Videnska 1083, 142 20, Prague, Czech Republic. petrmat@biomed.cas.cz
Hodny
Zdenek
Z
Svandova
Ivana
I
Svoboda
Petr
P
D018957:CD59 Antigens / Q000032:analysis* / Q000276:immunology; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D003902:Detergents / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis* / Q000276:immunology; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*
D018957:CD59 Antigens; D003902:Detergents; D018960:Hyaluronan Receptors; D008565:Membrane Proteins