ABCMdb

ABCC1 p.Lys684Met

[switch to full view]
Comments [show]

None has been submitted yet.

Publications

PMID: 10777615 [PubMed] Gao M et al: "Comparison of the functional characteristics of the nucleotide binding domains of multidrug resistance protein 1."

No. Sentence Comment

80 The primers with the mismatched bases (boldface type) for K684M and K1333M were 5Ј-GGCTGCGGAATGTCGTC- CCTG-3Ј and 5Ј-GGAGCTGGGATGTCGTCCCTG-3Ј, respectively. Login to comment
83 The Bsu36I-SphI fragment bearing the K684M mutation was isolated from pGEM-NBD1 and used to replace the same region in pFB-MRP1 (35) and pFBDual-halves to create pFB-MRP1/K684M and pFBDual-halves/ K684M, respectively. Login to comment
85 The SphI-KpnI fragment was then isolated from the resulting plasmid and used to replace the same region in pFB-MRP1 and pFB-MRP1/K684M to give pFB-MRP1/K1333M and pFB-MRP1/Double Km, respectively. Login to comment
87 Then the SalI-XbaI fragment of pFBDual-halves/K684M was isolated and cloned into the resulting vector as described for pFBDual-halves. Login to comment
204 The rate of ATP/ MRP1-dependent LTC4 uptake by vesicles from cells expressing the NBD1 mutant, MRP1/K684M, was approximately 25% (at 1 min and in the presence of 50 nM LTC4) of that obtained with vesicles containing the wild-type protein (Fig. 6B). Login to comment
217 Time and ATP-dependence of uptake is shown for MRP1 (f), NBD1 mutant MRP1/K684M (Œ), NBD2 mutant MRP1/K1333M (), and the double mutant MRP1/Double KM (ࡗ). Login to comment
226 Thus, despite the transport activity observed with the K684M mutant, no trapping of 8-azido-ADP was detectable. Login to comment
227 LTC4 Transport by Co-expressed Half-molecules of MRP1 Containing Walker A Lysine Mutations-To further characterize the effect of the Walker A K684M and K1333M substitutions on the ability to photolabel each NBD with 8-azido-␣- [32 P]ATP, these mutations were introduced into each of the half-molecules, which were then expressed either together, or with the appropriate wild-type half-molecule, using dual expression vectors. Login to comment
230 The rate of LTC4 uptake by vesicles containing the K684M mutation was approximately 35% (at 1 min and in the presence of 50 nM LTC4) of that obtained with vesicles containing both halves of the wild-type protein (Fig. 7B). Login to comment
234 Despite similar levels of the NH2-proximal half-molecules in the membrane vesicles used (Fig. 7A), labeling of the NH2-proximal half-molecule containing the K684M mutation (for both Dual-halves/K684M and Dual-halves/Double KM) was not detectable with either 8-azido-␣- (Fig. 8B) or 8-azido-␥-[32 P]ATP (Fig. 8A), regardless of whether it was expressed with a wild-type or mutant COOH-proximal half-molecule. Login to comment
235 In addition, the K684M mutation eliminated any LTC4 enhancement of the photolabeling of NBD1 and all labeling of a co-expressed wild-type COOH-proximal half-molecule. Login to comment
242 Consistent with the results obtained with the full-length protein, no labeling of either NBD was observed when either the K1333M or K684M mutant half-molecules were co-expressed with the appropriate wild-type half of the protein despite the demonstrable transport activity of the latter combination. Login to comment
247 A, membrane proteins from Sf21 cells expressing both halves of wild-type MRP1 (Dual-halves), NBD1 mutant (Dual-halves/ K684M), NBD2 mutant (Dual-halves/K1333M) and double mutant (Dual-halves/Double KM) were separated by SDS-PAGE on a 5-15% gradient gel and transferred to Immobilon-P membranes. Login to comment
250 Time and ATP dependence of uptake is shown for Dual-halves (f), Dual-halves/K684M (Œ), Dual-halves/K1333M (), and Dual-halves/ Double KM (ࡗ). Login to comment
255 Thus, the results of LTC4 transport studies were very similar to those obtained with K684M mutation. Login to comment
264 Thus, the effect of the Ins708 mutation was similar to that of the K684M mutation both with respect to LTC4 transport activity and labeling by 8-azido-ATP and -ADP. Login to comment
317 The K684M mutation had a similar effect on the activity of the reconstituted transporter, and we confirmed that binding of 8-azido-ATP by the mutated NBD1 had been abolished. Login to comment
318 However, despite the retention of 30% of the wild-type level of ATP-dependent transport activity, the K684M mutation also eliminated detectable trapping of 8-azido-ADP by NBD2. Login to comment
319 Consequently, to ensure that the transport activity detected in the K684M mutation was indeed dependent on ATP hydrolysis, assays were carried out with the nonhydrolyzable ATP analogue, ATP␥S, and no transport activity could be detected (data not shown). Login to comment
324 Furthermore, like the K684M mutation, it reduced LTC4 transport by approximately 70%. Login to comment
325 Thus, the Ins708 mutation behaved in a manner indistinguishable from the K684M mutation with respect to nucleotide binding, vanadate-induced trapping, and transport activity. Login to comment
326 No major differences could be detected between the ATP dependence of the initial rates of transport of the K684M and Ins708 mutations when compared with the wild-type protein. Login to comment
327 As observed with the wild-type protein, initial rates of LTC4 transport of both the K684M and Ins708 mutations reached a maximum between 0.5 and 1 mM ATP (data not shown). Login to comment
329 The inability to trap nucleotide at NBD2 in the K684M and the Ins708 mutant proteins suggests that in the absence of ATP binding and possibly hydrolysis at NBD1, either 8-azidoADP is released rapidly from NBD2 even in the presence of vanadate or that the conformation in which NBD2 binds the 8-azido-ADP vanadate complex cannot be efficiently photoaffinity-labeled. Login to comment
332 The combined results of LTC4 transport and photoaffinity labeling studies with the K684M, Ins708, and K1333M mutants are consistent with a model in which ATP hydrolysis at NBD1 is obligatorily coupled to hydrolysis at NBD2 but not vice versa. Login to comment
334 The lack of reciprocal coupling between the two NBDs of MRP1 and the different consequences of the K684M and K1333M mutations raise an important question with respect to the role played by NBD1 in substrate transport. Login to comment
336 However, it is not possible to determine with the data presently available for MRP1 whether the decrease in LTC4 transport efficiency seen with the K684M mutation is a direct consequence of the inactivation of NBD1 or the result of a decrease in the efficiency of ATP hydrolysis at NBD2. Login to comment
337 Several observations, including (i) the LTC4-dependent stimulation of 8-azido-ATP binding by NBD1, (ii) the retention of partial transport activity following inactivation of NBD1 but not NBD2 and, (iii) loss of the ability to trap and photolabel NBD2 in the K684M and Ins708 mutants with 8-azido-ADP, are equally compatible with a mechanism in which the role of NBD1 is to regulate, in a substrate-responsive manner, the efficiency of ATP binding and hydrolysis at NBD2. Login to comment

PMID: 11507101 [PubMed] Qian YM et al: "Characterization of binding of leukotriene C4 by human multidrug resistance protein 1: evidence of differential interactions with NH2- and COOH-proximal halves of the protein."

No. Sentence Comment

149 Consistent with the suggestion that the two NBDs of MRP1 fulfill different functional roles, our previous studies demonstrated that Walker A mutations, K684M and K1333M, in NBD1 and NBD2, respectively, had different effects on the ability of the protein to transport LTC4 (25). Login to comment
151 As shown in Fig. 8C, either the double mutation K684M/K1333M or the single mutation K1333M abolished the ability of vanadate and ATP to inhibit LTC4 binding, whereas mutation K684M did not. Login to comment
235 C, effect of vanadate trapping on [3 H]LTC4 labeling of co-expressed expressed mutant forms of MRP11-932 and MRP1932-1531 in which NBD1 or NBD2 had been in activated by mutation of essential Walker A mutations (K684M in NBD1 and K1333M in NBD). Login to comment

PMID: 15152943 [PubMed] Szentpetery Z et al: "Functional studies on the MRP1 multidrug transporter: characterization of ABC-signature mutant variants."

No. Sentence Comment

42 When a highly conserved lysine residue within the Walker A motif in MRP1 was substituted for methionine in either the N-ABC or in the C-ABC unit, the K1333M mutation in the C-ABC nearly abolished ATP-dependent LTC4 uptake, whereas the K684M substitution in the N-ABC had a less pronounced effect (22,23). Login to comment

PMID: 15155846 [PubMed] Ren XQ et al: "Function of the ABC signature sequences in the human multidrug resistance protein 1."

No. Sentence Comment

3 We therefore investigated the effect of mutation of the signature sequences (G771D and G1433D) and of the Walker A motifs (K684M and K1333M) in the NBDs on the 8-azido-[␣-32 P]ATP photolabeling and 8-azido-[␣-32 P]ADP vanadate trapping of MRP1. Login to comment
8 Trapping at both NBD1 and NBD2 was almost completely inhibited by K684M and K1333M mutations and by the K684M/K1333M double mutation. Login to comment
55 The MRP1 construct encoding the K684M mutation was generated by site-directed mutagenesis using the reverse primer 5Ј-GAGAGCAGGGACAGCATTCCG- CAGCCC-3Ј (bold indicates a mismatched base encoding the K684M mutation; underlining indicates a silent mutation). Login to comment
57 The MRP1 K684M/K1333M double mutant was generated by exchanging DNA fragments from the single mutations. Login to comment
98 Furthermore, G771D mutation in signature sequence of NBD1 more effectively lowered LTC4 uptake activity than K684M in Walker A motif of the same NBD, suggesting that the signature sequence has a more important role than the Walker A motif in the transport of the substrate. Login to comment
113 Mutation of the Walker A motif in the N-terminal (K684M) NBD1 or the C-terminal (K1333M) NBD2 almost completely inhibited the labeling of the NBD in their respective fragments. Login to comment
127 Either single or double mutation of the Walker A motifs in NBD1 and/or NBD2 (K684M, K1333M, or K684M/K1333M double mutations) almost completely inhibited trapping by both NBD1 and NBD2 domains (Fig. 7A). Login to comment
141 Although ATP-dependent LTC4 transport by G771D and G1433D MRP1 mutants, as well as transport by the K684M and K1333M mutants in the Walker A motifs, were considerably decreased, GSH-dependent photolabeling with azido AG-A of these MRP1 mutants was retained. Login to comment

PMID: 15755910 [PubMed] Payen L et al: "Functional interactions between nucleotide binding domains and leukotriene C4 binding sites of multidrug resistance protein 1 (ABCC1)."

No. Sentence Comment

127 Despite the retention of ϳ30% of wt LTC4 transport activity by the K684M mutant protein, we were unable to detect photolabeling of either NBD with 8-azido-ATP (Gao et al., 2000). Login to comment

PMID: 17295059 [PubMed] Chang XB et al: "A molecular understanding of ATP-dependent solute transport by multidrug resistance-associated protein MRP1."

No. Sentence Comment

256 Mutation of the Walker A motif K684 residue in NBD1, such as K684L [40, 141, 148], K684M [16, 63, 118], K684R [61] or K684E [61], significantly reduced ATP binding (at 4°C) at the mutated NBD1 and the intact NBD2 and Vi dependent ADP trapping at 37°C, but never completely abolished ATP-dependent solute transport. Login to comment