1 research outputs found
ABCB1 Does Not Require the Side-Chain Hydrogen-Bond Donors Gln347, Gln725, Gln990 to Confer Cellular Resistance to the Anticancer Drug Taxol
The multidrug efflux transporter ABCB1 is clinically important for drug absorption and
distribution and can be a determinant of chemotherapy failure. Recent structure data shows that three
glutamines donate hydrogen bonds to coordinate taxol in the drug binding pocket. This is consistent
with earlier drug structure-activity relationships that implicated the importance of hydrogen bonds
in drug recognition by ABCB1. By replacing the glutamines with alanines we have tested whether
any, or all, of Gln347, Gln725, and Gln990 are important for the transport of three different drug classes.
Flow cytometric transport assays show that Q347A and Q990A act synergistically to reduce transport
of Calcein-AM, BODIPY-verapamil, and OREGON GREEN-taxol bisacetate but the magnitude of
the effect was dependent on the test drug and no combination of mutations completely abrogated
function. Surprisingly, Q725A mutants generally improved transport of Calcein-AM and BODIPYverapamil, suggesting that engagement of the wild-type Gln725 in a hydrogen bond is inhibitory for
the transport mechanism. To test transport of unmodified taxol, stable expression of Q347/725A
and the triple mutant was engineered and shown to confer equivalent resistance to the drug as
the wild-type transporter, further indicating that none of these potential hydrogen bonds between
transporter and transport substrate are critical for the function of ABCB1. The implications of the
data for plasticity of the drug binding pocket are discussed