Drug-stimulated ATPase activity of a deletion mutant of the human multidrug-resistance protein (MDR1).
The baculovirus-insect cell system has been used for the functional expression of the human multidrug resistance protein (MDR1) and a mutant MDR1 variant lacking a twenty amino acid segment from the first extracellular loop (delta aa78-97 MDR1). Both MDR1 proteins were found to be correctly inserted into the insect cell membrane as indicated by their interaction with MRK 16 antibody. The removal of the 78-97 segment from the first extracellular loop dramatically altered drug-stimulated ATPase activity. Rhodamine 123 or vinblastine were not able to stimulate the mutant protein and Calcein AM had also little effect. In contrast, verapamil increased the ATPase activity of the mutant almost to the same maximal level as that of the wild type. However, the verapamil concentration needed for the half maximal stimulation of the ATPase activity was found to be about hundred times higher than that for the wild type MDR1. These results indicate that a partial deletion of an extracellular loop modulates the affinity of MDR1 for its transportable substrates in a variable fashion.
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000302:isolation & purification / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000302:isolation & purification / Q000378:metabolism*; D016367:Baculoviridae; D002462:Cell Membrane / Q000378:metabolism; D005434:Flow Cytometry; D014644:Genetic Variation; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D011994:Recombinant Proteins; D000251:Adenosine Triphosphatases