ABCMdb

PMID: 10455147

Loo TW, Clarke DM
The transmembrane domains of the human multidrug resistance P-glycoprotein are sufficient to mediate drug binding and trafficking to the cell surface.
J Biol Chem. 1999 Aug 27;274(35):24759-65., 1999-08-27 [PubMed]

Sentences

No. Mutations Sentence Comment

29 ABCB1 p.Pro709Gly EXPERIMENTAL PROCEDURES Generation of Deletion Mutants-The cDNAs coding for full-length MDR1 (36), mutant P709G (37), or the N-half and C-half P-gp molecules (14) and containing the epitope for monoclonal antibody A52 at the C-terminal ends were subcloned into the mammalian expression vector pMT21. Login to comment 43 ABCB1 p.Pro709Gly For expression studies involving drug resistance assays, the cDNAs of the A52-tagged wild-type, mutant P709G, N-half, C-half, and TMD1ϩ2 were also inserted into the Epstein-Barr virus-based vector, pREP4 (Invitrogen Inc.). Login to comment 137 ABCB1 p.Pro709Gly The cDNAs for the A52-tagged wild-type, and P-gp mutants P709G, N-half, C-half, and TMD1ϩ2 were inserted into the pREP4 vector and transfected into HEK 293(EBNA-1) cells. Login to comment 138 ABCB1 p.Pro709Gly Mutant P709G was included because it shows similar processing defects as mutant TMD1ϩ2. Login to comment 139 ABCB1 p.Pro709Gly When mutant P709G is expressed in HEK 293 cells in the absence of drug substrate, the major product is a protein of 150 kDa that is retained in the endoplasmic reticulum as a core-glycosylated intermediate (37). Login to comment 145 ABCB1 p.Pro709Gly For mutant P709G, the major product in the absence of drug substrate was the 150-kDa (core-glycosylated) protein, while the 170-kDa protein was the major product in the presence of substrate. When N-half (90-kDa) P-gp was co-expressed with C-half P-gp in the presence of substrate, another N-half protein of 115 kDa was present. Login to comment 155 ABCB1 p.Pro709Gly Fig. 6A shows that the mature forms of wild-type, mutant P709G, N-half, and TMD1ϩ2 P-gps were still present in the cells after treatment with cyclosporin A followed by vinblastine. Login to comment 170 ABCB1 p.Pro709Gly In mutant P709G or in the half-molecules of P-gp, preincubation of the cells with cyclosporin A greatly increased the relative resistance to vinblastine. Login to comment 171 ABCB1 p.Pro709Gly The cells expressing mutant P709G that were not pretreated with cyclosporin A showed about an 8-fold increase in resistance to vinblastine relative to the control cells. Login to comment 195 ABCB1 p.Pro709Gly A, HEK 293(EBNA-1) cells were transfected with pREP4 vector (B, Control) or pREP4 vectors containing cDNAs for full-length wild-type P-gp (Wild), mutant P709G, or TMD1ϩ2, or they were cotransfected with the cDNAs for the N-half and C-half of P-gp (Halves). Login to comment