ABCMdb

ABCB1 p.Ser909Cys

None has been submitted yet.

Publications

PMID: 17627029 [PubMed] Zolnerciks JK et al: "Evidence for a Sav1866-like architecture for the human multidrug transporter P-glycoprotein."

No. Sentence Comment

86 AGTTGTTTCTTTGAC-3Ј; and S909C, 5Ј-CCGAACCGTTG- TTTGTCTCACCCAGGAGCAGAAGTTTG-3Ј. Login to comment
139 The residues in ICL4 and NBD1 predicted to be in close proximity were replaced individually and also in the following pairwise combinations; Leu443Cys with Ser909Cys (L443CϩS909C), and Ser474Cys with Arg905Cys (S474CϩR905C). Login to comment
155 Each of the single cysteine substitution mutants L443C, S474C, R905C, and S909C retain significant levels of drug transport activity, as do the L443CϩS909C and S474CϩR905C double mutants (Fig. 3B, C). Login to comment
169 B) Histogram of cell surface expression of P-glycoproteins L443C, S474C, R905C, S909C and double mutants L443CϩS909C and S474CϩR905C are shown as indicated. Login to comment
173 However, to rule out the possibility that the gel retardation is the product of intermolecular cross linking between, for example, L443C on one P-glycoprotein with S909C on a second molecule, we cotransfected HEK293T cells with plasmids encoding both the single cysteine forms of the double mutants. Login to comment
180 Taken together with the results of the cross-linking, the simplest interpretation of these data is that L443C is in close proximity to S909C and that S474C is in close proximity to R905C in the tertiary structure of monomeric P-glycoprotein, entirely consistent with the hypothesis based on the crystal structure of Sav1866. Login to comment
183 Both treatments caused a reproducible reduction in the level of cross-linking between L443C and S909C (fewer slow migrating P-glycoprotein species and more noncrosslinked species) with each of the three chemical cross-linkers, although the effect seems more pronounced with BMH (Fig. 7A). Login to comment
207 Efflux activity of cysteine mutants wt Pgp-cys- L443C S909C L443CϩS909C S474C R905C S474CϩR905C E556Q Mock Mean % activity 100 100.8 100.7 99.9 100.6 99.5 96.9 101.2 8.6 0.0 (Ϯse) Ϯ 0.6 Ϯ 0.8 Ϯ 0.4 Ϯ 0.5 Ϯ 1.4 Ϯ1.7 Ϯ0.9 Ϯ1.2 Ϯ 11.9 Ϯ 8.9 n 6 6 2 2 4 2 4 2 6 4 The efflux activity of the mutants generated in this study is presented as a percentage of the drug efflux activity of wild-type P-glycoprotein (wt), as described in Materials and Methods. Login to comment
212 SDS-PAGE and Western analyses of P-glycoproteins with novel cysteines introduced into NBD1 (L443C and S474C) and/or ICL4 of TMD2 (R905C and S909C), as indicated. Login to comment
219 L443C P-glycoprotein was coexpressed in cells with S909C P-glycoprotein, and likewise S474C P-glycoprotein was coexpressed with R905C P-glycoprotein. Login to comment
220 SDS-PAGE and Western analyses showed no evidence of intermolecular cross-linking (lack of a slow mobility P-glycoprotein species) between the L443C on one protein and S909C on another, or between S474C and R905C on different P-glycoproteins. Login to comment
245 SDS-PAGE and Western analyses of P-glycoprotein double mutants with novel cysteines introduced at (A) L443C and S909C, or (B) S474C and R905C. Login to comment
257 Evidence in support of the interaction between ICL4 and NBD1 being a site of energy transduction through which the ATP and drug binding sites are coupled allosterically is provided by the modulation of the cross-linking efficiency between L443C and S909C by the nonhydrolysable ATP analog AMP-PNP, and also by ADP in the presence of vanadate (ADP.Vi) (Fig. 7A). Login to comment
264 While it is unlikely that all the P-glycoprotein molecules in a membrane fraction would be accessible to nucleotide (the NBDs of some in the fraction are likely to be intravesicular), the data suggest that in the presence of AMP-PNP or ADP.Vi the L443C and S909C cysteines are less available for cross-linking. Login to comment

PMID: 22700974 [PubMed] Loo TW et al: "The ATPase activity of the P-glycoprotein drug pump is highly activated when the N-terminal and central regions of the nucleotide-binding domains are linked closely together."

No. Sentence Comment

106 The location of residues that were mutated to cysteine to test for the effect of cross-linking between NBD1 and NBD2 (P517C/I1050C), NBD1 and TMD2 (L443C/S909C), ICL1 and ICL3 (D177C/N820C), TM segments 2 and 11 (C137/A935C) or 6 and 12 (T333C/L975C) are shown. Login to comment
152 The ATPase activity of M17M cross-linked L175C/N820C could still be highly activated when assayed in the presence of verapamil. Login to comment
154 ATPase Activity Is Not Activated when the Homologous Halves Are Cross-linked at Locations Predicted Not to Undergo Large Distance Changes during the Open to Closed Conformational Change-Mutant C137/A935C contains natural Cys-137 in TM segment 2 (TMD1) and the A935C mutation in TM segment 11 (TMD2). Login to comment
156 Mutant L443C/S909C contains a cysteine (L443) in NBD1 and a cysteine (S909C) in the ICL4 loop that connects the cytoplasmic extensions of TM segments 10 and 11. Login to comment
158 In previous studies we showed that mutants C137/A935C (39) and L443C/ S909C (27) had verapamil-stimulated ATPase activities similar to Cys-less P-gp. Login to comment
159 Membranes prepared from cells expressing mutants C137/ A935C or L443C/S909C were treated with M4M cross-linker. Login to comment
163 These results indicate that stimulation of ATPase activity observed in M4M cross-linked mutants P517C/I1050C (Fig. 2B), D177C/N820C, or L175C/N820C (Fig. 3C) was likely a specific effect of trapping the protein in the closed conformation. Login to comment
166 A, membranes prepared from HEK 293 cells expressing the NBD1/TMD2 mutant L443C/ S909C or the TM2/TM11 mutant C137/A935C were treated with (ϩ) or without (-) the M4M cross-linker. Login to comment
168 The positions of the cross-linked (X-link) and mature (170 kDa) P-gps are indicated. B, membranes expressing mutants L443C/ S909C or C137/A935C were treated with or without (None) M4M cross-linker and histidine-tagged P-gp isolated by nickel-chelate chromatography. Login to comment
287 One explanation is that some movement is required at the C137/A935C and L443C/S909C interfaces during the catalytic cycle. Login to comment
301 Direct cross-linking of cysteines in the mutants would trap C137/A935C or L443C/S909C at a distance of about 5.6 Å, the average distance between ␣ carbons in a disulfide bond (64). Login to comment
304 Another plausible explanation as to why direct cross-linking of mutants C137/A935C or L443C/S909C inhibited activity is that cross-linking may have trapped the protein in an inactive conformation with the sites closer than normal. Login to comment
104 The location of residues that were mutated to cysteine to test for the effect of cross-linking between NBD1 and NBD2 (P517C/I1050C), NBD1 and TMD2 (L443C/S909C), ICL1 and ICL3 (D177C/N820C), TM segments 2 and 11 (C137/A935C) or 6 and 12 (T333C/L975C) are shown. Login to comment
161 A, membranes prepared from HEK 293 cells expressing the NBD1/TMD2 mutant L443C/ S909C or the TM2/TM11 mutant C137/A935C were treated with (af9;) or without (afa;) the M4M cross-linker. Login to comment
280 One explanation is that some movement is required at the C137/A935C and L443C/S909C interfaces during the catalytic cycle. Login to comment
294 Direct cross-linking of cysteines in the mutants would trap C137/A935C or L443C/S909C at a distance of about 5.6 &#c5;, the average distance between ॷ carbons in a disulfide bond (64). Login to comment
297 Another plausible explanation as to why direct cross-linking of mutants C137/A935C or L443C/S909C inhibited activity is that cross-linking may have trapped the protein in an inactive conformation with the sites closer than normal. Login to comment