The role of hydrogen bond acceptor groups in the interaction of substrates with Pdr5p, a major yeast drug transporter.
The yeast ABC (ATP-binding cassette protein) multidrug transporter Pdr5p transports a broad spectrum of xenobiotic compounds, including antifungal and antitumor agents. Previously, we demonstrated that substrate size is an important factor in substrate-transporter interaction and that Pdr5p has at least three substrate-binding sites. In this study, we use a combination of whole cell transport assays and photoaffinity labeling of Pdr5p with [(125)I]iodoarylazidoprazosin in purified plasma membrane vesicles to study the behavior of two series of novel substrates: trityl (triphenylmethyl) and carbazole derivatives. The results indicate that site 2, defined initially by tritylimidazole efflux, requires at least a single hydrogen bond acceptor group (electron pair donor). In contrast, complete inhibition of rhodamine 6G efflux and [(125)I]iodoarylazidoprazosin binding at site 1 requires substrates with three electronegative groups. Carbazole and trityl substrates with two groups show saturating, incomplete inhibition at this site. This type of inhibition is frequently observed in bacterial multidrug-binding proteins that use a pocket with multiple binding sites. The presence of multiple sites with different requirements for substrate-Pdr5p interaction may explain the broad specificity of xenobiotic compounds transported by this protein.
Hanson
Leanne
L
Department of Biology, The Catholic University of America, Washington, DC 20064, USA.
May
Leopold
L
Tuma
Pamela
P
Keeven
James
J
Mehl
Patrick
P
Ferenz
Michelle
M
Ambudkar
Suresh V
SV
Golin
John
J
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000378:metabolism*; D000935:Antifungal Agents / Q000378:metabolism; D000970:Antineoplastic Agents / Q000378:metabolism; D001386:Azides / Q000378:metabolism; D001665:Binding Sites / Q000187:drug effects; D001692:Biological Transport / Q000187:drug effects; D002227:Carbazoles / Q000737:chemistry / Q000378:metabolism; D003022:Clotrimazole / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D004611:Ellipticines / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding / Q000187:drug effects; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000737:chemistry / Q000378:metabolism; D011224:Prazosin / Q000031:analogs & derivatives / Q000378:metabolism; D012235:Rhodamines / Q000037:antagonists & inhibitors / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D013379:Substrate Specificity / Q000187:drug effects; D014316:Tritium; D014320:Triphenylmethyl Compounds / Q000737:chemistry / Q000378:metabolism; D015262:Xenobiotics / Q000737:chemistry / Q000378:metabolism*
D018528:ATP-Binding Cassette Transporters; D000935:Antifungal Agents; D000970:Antineoplastic Agents; D001386:Azides; D002227:Carbazoles; D003022:Clotrimazole; D003432:Cross-Linking Reagents; D004611:Ellipticines; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011224:Prazosin; D012235:Rhodamines; D029701:Saccharomyces cerevisiae Proteins; D014316:Tritium; D014320:Triphenylmethyl Compounds; D015262:Xenobiotics; C098803:1-trityl-4-nitroimidazole; C092438:PDR5 protein, S cerevisiae; C026188:rhodamine 6G; C041514:carbazole; C034192:ellipticine; C020702:3-amino-9-ethylcarbazole; C041552:azidoprazosin