Molecular study of P-glycoprotein in multidrug resistance using surface plasmon resonance.
P-Glycoprotein is an integral membrane protein which mediates the energy-dependent efflux of various antitumor agents from multidrug-resistant cancer cells. Surface plasmon resonance was used for the detection of P-glycoprotein after solubilization from drug-resistant and drug-sensitive Chinese hamster ovary cells and for the analysis of its interaction with cyclosporin A, a competitive inhibitor of drug efflux. Detection of P-glycoprotein relied on its binding to the monoclonal antibody C219 which was immobilized on a sensor chip. Binding of Zwittergent 3-14-solubilized P-glycoprotein to the antibody was concentration-dependent and reflected the relative abundance of P-glycoprotein in both cell lines. It was abolished when C219 was omitted or replaced by a rabbit anti-mouse IgG antibody and considerably reduced after precipitation of P-glycoprotein with wheat germ agglutinin. Preincubation of solubilized proteins with cyclosporin A increased the amount of protein bound to the antibody by approximately 30%. These results indicate that surface plasmon resonance is well suited to the detection of P-glycoprotein from biological samples and shows promise as a tool for the study of its interaction with different drugs.
Demeule
M
M
Laboratoire de membranologie, Université du Québec à Montréal, Canada.
Vachon
V
V
Delisle
M C
MC
Beaulieu
E
E
Averill-Bates
D
D
Murphy
G F
GF
Béliveau
R
R
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis* / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D015374:Biosensing Techniques; D016466:CHO Cells; D006224:Cricetinae; D016572:Cyclosporine / Q000378:metabolism; D018432:Drug Resistance, Multiple; D007700:Kinetics; D051379:Mice; D011817:Rabbits; D012995:Solubility
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000911:Antibodies, Monoclonal; D016572:Cyclosporine