posted on 2016-02-19, 14:27authored byPetra Kueppers, Rakeshkumar
P. Gupta, Jan Stindt, Sander
H. J. Smits, Lutz Schmitt
The pleiotropic drug resistance network
in budding yeast presents
a first line of defense against xenobiotics, which is formed by primary
and secondary active membrane transporters. Among these transporters,
the ABC transporter Pdr5 is a key component, because it confers resistance
against a broad spectrum of such cytotoxic agents. Furthermore, it
represents a model system for homologous transporters from pathogenic
fungi and has been intensively studied in the past. In addition to
other mutational studies, the S1360F mutation of Pdr5 was found to
modulate substrate specificity and resistance. Notably, in the S1360F
background, the resistance against the immunosuppressant FK506 is
drastically increased. We present a detailed analysis of this mutation
that is located in the predicted cytosolic part of transmembrane helix
11. Our data demonstrate that kinetic and thermodynamic parameters
of the S1360F mutant are similar to those of the wild-type protein,
except for FK506-inhibited ATPase activity and the degree of competitive
inhibition. In summary, our results indicate that the S1360F mutation
within the transmembrane domain interferes drastically with the ability
of the nucleotide-binding domains to hydrolyze ATP by interfering
with interdomain crosstalk.