ABCMdb

ABCB1 p.Ala729Cys

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Publications

PMID: 16813563 [PubMed] Loo TW et al: "Transmembrane segment 7 of human P-glycoprotein forms part of the drug-binding pocket."

No. Sentence Comment

74 Mutant Verapamil S50 (µM) Vinblastine S50 (µM) Colchicine S50 (µM) F711C 9.8 + - 1.1 2.3 + - 0.2 410 + - 20 V712C 10.0 + - 0.8 1.9 + - 0.2 320 + - 40 V713C 9.9 + - 1.3 1.8 + - 0.2 340 + - 40 G714C 14.1 + - 2.1 1.9 + - 0.3 410 + - 30 V715C 9.8 + - 1.3 1.8 + - 0.3 330 + - 40 F716C 10.1 + - 1.0 2.1 + - 0.1 340 + - 30 C717C 10.0 + - 1.5 2.3 + - 0.3 390 + - 20 A718C 10.5 + - 1.3 2.0 + - 0.1 330 + - 30 I719C 10.5 + - 1.5 1.9 + - 0.2 370 + - 30 I720C 12.4 + - 0.7 2.4 + - 0.2 390 + - 10 N721C 13.0 + - 1.2 1.7 + - 0.2 380 + - 40 G722C ND ND ND G723C 11.1 + - 0.4 2.0 + - 0.3 410 + - 20 L724C 12.0 + - 1.5 2.4 + - 0.3 340 + - 30 Q725C 11.7 + - 1.1 2.0 + - 0.1 390 + - 20 P726C 11.9 + - 1.0 1.9 + - 0.2 450 + - 30 A727C 21.2 + - 3.1 1.9 + - 0.2 480 + - 40 F728C 48.7 + - 2.2 2.7 + - 0.3 830 + - 90 A729C 34.3 + - 3.0 2.3 + - 0.2 760 + - 80 I730C 12.0 + - 1.1 2.0 + - 0.1 420 + - 30 I731C 13.5 + - 1.9 1.9 + - 0.2 410 + - 30 Cys-less 11.6 + - 1.3 2.1 + - 0.3 400 + - 40 into the endoplasmic reticulum during synthesis of the protein. Login to comment
85 Table 1 shows that two mutants, F728C and A729C, showed about 4.2and 3-fold decreases respectively, in the apparent affinity for verapamil when compared with Cys-less P-gp. Login to comment
97 Samples of the isolated P-gp were mixed with lipid and assayed for ATPase activity and compared with untreated P-gp. All of the mutants, except Q725C, P726C, F728C and A729C, were unaffected by treatment with MTS-verapamil (Figure 2A). Login to comment
99 By contrast, the ATPase activity of mutants Q725C, F728C and A729C increased 2.6-, 4.7and 4.5-fold respectively, compared with untreated mutant P-gp. Login to comment
100 Although mutants Q725C, F728C and A729C showed increased ATPase activity after treatment with MTS-verapamil, the activities were less than 50% of the verapamil-stimulated ATPase activity of Cys-less P-gp (maximum 11.1-fold stimulation; Figure 2B). Login to comment
101 We have shown previously that mutants Q725C, F728C and A729C were stimulated more than 10-fold with verapamil [37]. Login to comment
103 Therefore it was possible that reaction of MTS-verapamil with mutants Q725C, F728C and A729C was incomplete or that they were completely modified but the bound verapamil was in a sub-optimal conformation for activation of ATPase activity. Login to comment
104 To distinguish between these two possibilities, mutants Q725C, F728C and A729C (activated by 0.3 mM MTS-verapamil) were treated with or without 3 mM MTS-verapamil. Login to comment
106 Figure 2(B) shows that increasing the level of MTS-verapamil from 0.3 mM to 3 mM did not decrease or increase the basal ATPase activity of mutant A729C. Login to comment
107 In addition, the activity of MTS-verapamil- modified mutant A729C could not be activated further by unmodified verapamil (Figure 2B). Login to comment
108 Therefore MTS-verapamil modification of mutant A729C inhibits its verapamil-stimulated ATPase activity. Login to comment
112 (B) Cys-less, Q725C, F728C and A729C P-gp mutants were treated with or without 3 mM MTS-verapamil and His-tagged P-gp isolated by nickel-chelate chromatography. Equivalent amounts of P-gp were mixed with lipid and ATPase activity determined in the presence (+) or absence (-) of 1 mM verapamil (Ver). Login to comment