ABCMdb

PMID: 16554024

Ohashi-Kobayashi A, Ohashi K, Du WB, Omote H, Nakamoto R, Al-Shawi M, Maeda M
Examination of drug resistance activity of human TAP-like (ABCB9) expressed in yeast.
Biochem Biophys Res Commun. 2006 May 5;343(2):597-601. Epub 2006 Mar 10., [PubMed]

Sentences

No. Mutations Sentence Comment

23 ABCB9 p.Lys545Met For YEpTAPL(K546M)His, the DraIII-NgoMI fragment from phTAPL(K545M) plasmid, which has a mutation of Lys (AAG) at position 546 to Met (AtG) by PCR technique reported by Imai et al. [11] using mutated primers, was excised and replaced by the DraIII-NgoMI fragment of YEpTAPLHis. Login to comment 82 ABCB9 p.Lys545Met For this purpose, we constructed an expression plasmid for hTAPL(K545M) harboring mutation in the Walker A motif, and it was introduced into yeast. Login to comment 83 ABCB9 p.Lys545Met ABCB9 p.Lys545Met For this purpose, we constructed an expression plasmid for hTAPL(K545M) harboring mutation in the Walker A motif, and it was introduced into yeast. Login to comment 84 ABCB9 p.Lys545Met ABCB9 p.Lys545Met Prior to drug sensitivity assay, we determined the ATP-binding capabilities of hTAPL and hTAPL(K545M). Login to comment 85 ABCB9 p.Lys545Met Expression of hTAPL(K545M) was similar in amounts compared with that of hTAPL by means of Western blotting. Login to comment 86 ABCB9 p.Lys545Met 3A and B, hTAPL bound ATP-agarose, whereas hTAPL(K545M) did not. Login to comment 87 ABCB9 p.Lys545Met 3A and B, hTAPL bound ATP-agarose, whereas hTAPL(K545M) did not. Login to comment 93 ABCB9 p.Lys545Met ABCB9 p.Lys545Met TAPL p.c. P P S S - + - + (ATP) - + - + (ATP) TAPL K545M p.c. P P S S hTAPL(K545M) Raf61 hMDR hTAPL 5 10 3 1 10 3 2 10 2 4 10 1 8 10 0 100&#b5;M valinomycin (cells/&#b5;l) A B Fig. 3. Login to comment 94 ABCB9 p.Lys545Met ABCB9 p.Lys545Met TAPL p.c. P P S S - + - + (ATP) - + - + (ATP) TAPL K545M p.c. P P S S hTAPL(K545M) Raf61 hMDR hTAPL 5103 1103 2102 4101 8100 100µM valinomycin (cells/µl) A B Fig. 3. Login to comment 95 ABCB9 p.Lys545Met (A) ATP-binding activity was examined on hTAPL and hTAPL(K545M) (mutation in Walker A motif). Login to comment 96 ABCB9 p.Lys545Met ABCB9 p.Lys545Met (A) ATP-binding activity was examined on hTAPL and hTAPL(K545M) (mutation in Walker A motif). Login to comment 97 ABCB9 p.Lys545Met Plasmid containing hTAPL or hTAPL(K545M) was transfected to yeast and the membrane fractions were prepared. Login to comment 100 ABCB9 p.Lys545Met (B) Sensitivity to valinomycin was examined on the yeast cells expressing hTAPL, hTAPL(K545M), or carrying control empty vector. Login to comment 101 ABCB9 p.Lys545Met (B) Sensitivity to valinomycin was examined on the yeast cells expressing hTAPL, hTAPL(K545M), or carrying control empty vector. Login to comment 102 ABCB9 p.Lys545Met Next, we examined valinomycin sensitivity of the yeast cell expressing hTAPL(K545M). Login to comment 103 ABCB9 p.Lys545Met Next, we examined valinomycin sensitivity of the yeast cell expressing hTAPL(K545M). Login to comment