title: Photoaffinity labeling under non-energized conditions of a specific drug-binding site of the ABC multidrug transporter LmrA from Lactococcus lactis.
abstract: The Lactococcus lactis multidrug resistance ABC transporter protein LmrA has been shown to confer resistance to structurally and functionally diverse antibiotics and anti-cancer drugs. Using a previously characterized photoreactive drug analogue of Rhodamine 123 (iodo-aryl azido-Rhodamine 123 or IAARh123), direct and specific photoaffinity labeling of LmrA in enriched membrane vesicles could be achieved under non-energized conditions. This photoaffinity labeling of LmrA occurs at a physiologically relevant site as it was inhibited by molar excess of ethidium bromide>Rhodamine 6G>vinblastine>doxorubicin>MK571 (a quinoline-based drug) while colchicine had no effect. The MDR-reversing agents PSC 833 and cyclosporin A were similarly effective in inhibiting IAARh123 photolabeling of LmrA and P-glycoprotein. In-gel digestion with Staphyloccocus aureus V8 protease of IAARh123-photolabeled LmrA revealed several IAARh123 labeled polypeptides, in addition to a 6.8kDa polypeptide that comprises the last two transmembrane domains of LmrA.
authors: Institute of Parasitology, McGill University, Macdonald Campus, Ste-Anne de Bellevue, Que, Canada.
authors: Omar
authors: O
authors: Alqawi
authors: Gerrit
authors: G
authors: Poelarends
authors: Wil N
authors: WN
authors: Konings
authors: Elias
authors: E
authors: Georges
authors: Omar
authors: O
authors: Alqwai
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry*; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D003078:Colchicine / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000494:pharmacology; D018432:Drug Resistance, Multiple; D004996:Ethidium / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000494:pharmacology; D007364:Intercalating Agents / Q000494:pharmacology; D013294:Lactococcus lactis / Q000378:metabolism*; D008027:Light; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011233:Precipitin Tests; D011422:Propionates / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011804:Quinolines / Q000494:pharmacology; D012235:Rhodamines / Q000494:pharmacology; D014747:Vinblastine / Q000494:pharmacology
chemical_list: D018528:ATP-Binding Cassette Transporters; D000903:Antibiotics, Antineoplastic; D000972:Antineoplastic Agents, Phytogenic; D001426:Bacterial Proteins; D003078:Colchicine; D018076:DNA, Complementary; D004317:Doxorubicin; D004996:Ethidium; D005456:Fluorescent Dyes; D007364:Intercalating Agents; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D010455:Peptides; D011422:Propionates; D011804:Quinolines; D012235:Rhodamines; D014747:Vinblastine; C101824:LmrA protein, Lactococcus lactis; C026188:rhodamine 6G; C059141:verlukast