ABCMdb

ABCC1 p.Glu1455Asp

Predicted by SNAP2:	A: D (95%), C: D (95%), D: D (95%), F: D (95%), G: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (95%), M: D (95%), N: D (95%), P: D (95%), Q: D (95%), R: D (95%), S: D (95%), T: D (95%), V: D (95%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	A: D, C: D, D: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D,

[switch to compact view]
Comments [show]

None has been submitted yet.

Publications
[hide] Role of carboxylate residues adjacent to the conse... J Biol Chem. 2003 Oct 3;278(40):38537-47. Epub 2003 Jul 27. Payen LF, Gao M, Westlake CJ, Cole SP, Deeley RG
Role of carboxylate residues adjacent to the conserved core Walker B motifs in the catalytic cycle of multidrug resistance protein 1 (ABCC1).
J Biol Chem. 2003 Oct 3;278(40):38537-47. Epub 2003 Jul 27., 2003-10-03 [PMID:12882957]

Abstract [show]
MRP1 belongs to subfamily "C" of the ABC transporter superfamily. The nucleotide-binding domains (NBDs) of the C family members are relatively divergent compared with many ABC proteins. They also differ in their ability to bind and hydrolyze ATP. In MRP1, NBD1 binds ATP with high affinity, whereas NBD2 is hydrolytically more active. Furthermore, ATP binding and/or hydrolysis by NBD2 of MRP1, but not NBD1, is required for MRP1 to shift from a high to low affinity substrate binding state. Little is known of the structural basis for these functional differences. One minor structural difference between NBDs is the presence of Asp COOH-terminal to the conserved core Walker B motif in NBD1, rather than the more commonly found Glu present in NBD2. We show that the presence of Asp or Glu following the Walker B motif profoundly affects the ability of the NBDs to bind, hydrolyze, and release nucleotide. An Asp to Glu mutation in NBD1 enhances its hydrolytic capacity and affinity for ADP but markedly decreases transport activity. In contrast, mutations that eliminate the negative charge of the Asp side chain have little effect. The decrease in transport caused by the Asp to Glu mutation in NBD1 is associated with an inability of MRP1 to shift from high to low affinity substrate binding states. In contrast, mutation of Glu to Asp markedly increases the affinity of NBD2 for ATP while decreasing its ability to hydrolyze ATP and to release ADP. This mutation eliminates transport activity but potentiates the conversion from a high to low affinity binding state in the presence of nucleotide. These observations are discussed in the context of catalytic models proposed for MRP1 and other ABC drug transport proteins.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
69 The forward primers for D793E and E1455D were 5Ј-CCTCTTCGATGAGCCCCTCTCAGC-3Ј and 5Ј-CTTGTGTTG- GATGATGCCACGGCAGC-3Ј, respectively. Login to comment
72 The EcoRI-KpnI fragments with mutations at NBD2 were isolated from pGEM-NBD2 and used to replace the equivalent region in pBS-Asp45 to generate pBS-D45/E1455D. Login to comment
73 This was then digested with NcoI and KpnI and the NcoI-KpnI fragment was used to replace the equivalent region of pFBDual-Asp45 to give pFBDual-D45/E1455D. Login to comment
74 Finally, the SalI-XbaI fragment of pFBDual-halves was isolated and cloned into pFBDual-D45/E1455D, which had been digested with the same enzymes, to generate pFBDual-halves/E1455D. Login to comment
75 In the D793E/E1455D double mutant, the SalI-XbaI fragment with the mutation in NBD1 was isolated and ligated to pFBDual-D45/E1455D, which had been digested with the same enzymes, to generate pFBDual-halves/D793E/E1455D. Login to comment
122 Effect of D793E and E1455D Single and Double Mutations on LTC4 Transport Activity-LTC4 uptake by membrane vesicles from Sf21 cells expressing wild-type and mutant MRP1 half-molecules was determined at 23 °C, as described by Loe et al. (7). Login to comment
125 The transport activity of vesicles containing either the E1455D mutant fragment and the wild-type NH2-terminal half, or co-expressed D793E and E1455D mutant fragments was similar to that of the beta-Gus control (Fig. 2B). Login to comment
126 Thus the effects of the D793E and E1455D mutations on LTC4 transport were similar to previously described mutations of the Walker A motif that eliminate ATP hydrolysis and decrease ATP binding by NBD1 and NBD2, respectively (24, 25). Login to comment
127 Photolabeling with 8-Azido-[␥-32 P]ATP in the Presence of AMP-PNP at 4 °C-To determine whether or not the D793E and E1455D mutations altered ATP binding, studies were carried out at 4 °C using the radioactive photoactivable analog of ATP, 8-azido-[␥-32 P]ATP, hydrolysis of which results in loss of the ␥-32 P label (24, 33). Login to comment
130 ATP labeling of wild-type MRP1, D793E, E1455D single mutant proteins was similar and occurred preferentially at NBD1, whereas it was slightly decreased at NBD1 of the double mutant protein. Login to comment
132 Labeling at NBD2 in the E1455D and D793E/E1455D mutant proteins was strongly and moderately enhanced, respectively, relative to the wild-type NBDs. Login to comment
134 Nucleotide Trapping by Wild-type MRP1 and D793E, E1455D Single and Double Mutant Proteins Using 8-Azido-[␣- 32 P]ATP-To determine whether ATP hydrolysis by the mutant NBDs was altered, ADP trapping experiments were performed at 37 °C using various concentrations of 8-azido-[␣- 32 P]ATP in the presence or absence of vanadate. Login to comment
140 Increased labeling of the mutant NBD1 was observed at all 8-azido-[␣-32 P]ATP concentrations tested and unlike the wild-type NBD1, was readily detectable at 2.5 ␮M 8-azido-[␣-32 P]ATP with both the D793E and D793E/E1455D mutants (Fig. 3, A and C). Login to comment
143 Effect of D793E and E1455D single and double mutations on ATP-dependent LTC4 transport activity (B) and on ATP binding at 4 °C (C). Login to comment
144 A, membrane proteins (1 ␮g from Sf21 cells expressing both halves of either MRP1 (MRP1 dh), mutant proteins (D793E, E1455D, and D793E/E1455D)) were separated by SDS-PAGE on gradient gels and transferred to Immobilon-P membranes. Login to comment
148 B, membrane vesicles (2 ␮g) containing wild-type MRP1 (f), MRP1 mutants D793E (Œ), E1455D (), D793E/E1455D (q), and control beta-Gus vector (ࡗ) were assayed for ATP-dependent LTC4 transport activity at 23 °C for up to 3 min in transport buffer containing [3 H]LTC4 (50 nM, 0.13 ␮Ci), as described under "Experimental Procedures." Login to comment
151 C, at 4 °C, 8-azido-[␥-32 P]ATP photolabeling by wild-type MRP1 and mutant D793E, E1455D, and D793E/E1455D was carried out. Login to comment
156 Catalytic Cycle of MRP138540 The E1455D mutation dramatically increased photolabeling of the mutant NBD2 and unlike the D793E mutation, increased rather than decreased labeling of the co-expressed wild-type NBD1. Login to comment
159 Despite the striking increase in photolabeling of the E1455D mutant NBD2, in the double D793E/E1455D mutant, labeling occurred predominantly at NBD1 and was markedly diminished at NBD2 relative to the E1455D single mutant. Login to comment
161 Photolabeling with 8-Azido-[␥-32 P]ATP or 8-Azido-[␣-32 P]ADP at 37 °C-The relatively weak vanadate dependence of photolabeling observed when using 8-azido-[␣-32 P]ATP, particularly with the E1455D and D793E/E1455D mutants, raised the possibility that the mutant NBD2 may be capable of tight binding of both ATP and ADP. Login to comment
165 In contrast, both NBDs of the E1455D mutant were strongly labeled by 8-azido-[␥-32 P]ATP, while in the D793E/E1455D double mutant labeling of NBD2 was much reduced and labeling of NBD1 was essentially eliminated (Fig. 4A). Login to comment
166 In contrast to the ATP binding observed at 4 °C, photolabeling at 37 °C of the E1455D mutant NBD2 was stronger than the co-expressed wild-type NBD1 and was only slightly competed by 1 mM AMP-PNP. Login to comment
167 These experiments suggest that the tight binding of ATP by the E1455D mutant NBD2 stimulates the binding of ATP by the co-expressed wild-type NBD1 and conversely, that the increased trapping of ADP at the D793E mutant NBD1, diminishes ATP binding by both the co-expressed wild-type and E1455D mutant NBD2. Login to comment
169 Nucleotide trapping by D793E, E1455D single and double mutant proteins using 8-azido-[␣-32 P]ATP. Login to comment
170 A-C, at 37 °C, under hydrolytic conditions, the effect of the 8-azido-[␣-32 P]ATP concentration on ADP trapping by wild-type MRP1 and D793E (A), E1455D (B), and D793E/E1455D (C) mutant proteins was evaluated. Membrane vesicles (20 ␮g) were incubated with 8-azido-[␣-32 P]ATP (1-15 ␮M) in the absence (-) or presence (ϩ) of 1 mM vanadate for 15 min in transport buffer containing 5 mM MgCl2. Login to comment
176 Effect of D793E, E1455D single and double mutations on ATP binding and ADP labeling under hydrolytic conditions. Login to comment
177 A, at 37 °C, 8-azido-[␥-32 P]ATP photolabeling by wild-type MRP1 and D793E, E1455D single and double mutations was carried out. Login to comment
181 The position of the labeled MRP1 NH2-half and COOH-half polypeptides are indicated, and endogenous proteins labeled are indicated by E followed by arrows. B, at 37 °C, 8-azido-[␣- 32 P]ADP photolabeling by wild-type MRP1 and mutant D793E, E1455D, and D793E/E1455D proteins was evaluated. Login to comment
192 ADP binding by the E1455D single mutant was strongly increased at both NBDs relative to the co-expressed wild-type fragments although the majority of photolabeling occurred at the mutant NBD2. Login to comment
193 In the D793E/E1455D double mutant, photolabeling of NBD2 was diminished and labeling of NBD1 was increased relative to the single D793E and E1455D single mutants, so that ADP labeling was similar at both NBDs (Fig. 4B, lanes 2-9). Login to comment
194 Thus as observed with 8-azido-[␥-32 P]ATP, the increased binding of ADP by the E1455D mutant NBD2 appears to stimulate tight binding of ADP at both the wild-type and D793E mutant NBD1. Login to comment
195 In contrast, the increased ADP binding of the D793E mutant NBD1 decreases ADP binding by both the wild-type and E1455D mutant NBD2, as would be predicted by an alternating site model of catalysis. Login to comment
196 Evaluation of ADP Release by Wild-type and D793E, E1455D Single and Double Mutant Proteins-It has been demonstrated that ADP release constitutes the rate-limiting step in the normal catalytic cycle of P-GP (41). Login to comment
200 Consequently, we investigated the influence of D793E, E1455D, and D793E/ E1455D MRP1 mutants on ADP release by each NBD. Login to comment
205 As described above, weak photolabeling of NBD1 in samples cross-linked immediately after washing could be detected with the D793E and D793E/E1455D mutants (Fig. 5, lanes 3 and 5). Login to comment
207 In contrast, the E1455D mutation displayed very strong labeling of the mutant NBD2 that persisted following the reincubation period (Fig. 5, lanes 4 and 10). Login to comment
209 Unlike the single E1455D mutant, very little photolabeling of NBD2 was retained following reincubation of the D793E/E1455D double mutant (Fig. 5, lanes 5 and 11). Login to comment
210 To further investigate the impairment of ADP release by the E1455D and the D793E/E1455D mutants, cold ADP (1 mM) was added 3 min before the end of the initial nucleotide labeling with 8-azido-[␣-32 P]ATP (Fig. 5, lanes 6-8). Login to comment
211 ADP competed completely for 8-azido-␣-32 P-nucleotide trapping at both NBDs of the D793E mutant and at NBD1 of the D793E/E1455D double mutant indicating that the bound nucleotide was readily exchangeable (Fig. 5, lanes 6 and 8). Login to comment
212 However, relatively strong photolabeling of both NBDs was retained in the E1455D mutant. Login to comment
213 In the D793E/E1455D double mutant all labeling at NBD1 was lost and labeling of NBD2 was markedly decreased compared with the E1455D single mutant (Fig. 5, lanes 7 and 8). Login to comment
215 Thus the results indicate that the release of ADP by the E1455D NBD2 is severely impaired and that this also decreases the ability of the co-expressed wild-type NBD1, but not the D793E mutant, to exchange nucleotide. Login to comment
216 LTC4 Photolabeling by Wild-type and D793E, E1455D Single, and D793E/E1455D Double Mutant Proteins Under Hydrolytic and Non-hydrolytic Conditions-We have shown previously that LTC4 can photolabel MRP1 at sites in MSD2 and MSD3 and that photolabeling, particularly of the site in MSD2, is strongly attenuated under ADP trapping conditions. Login to comment
219 To examine this prediction further, we investigated the effect of the D793E and E1455D mutations on the binding of LTC4 because the former, unlike the Walker A mutations, increases ADP trapping at NBD1, whereas the latter increases nucleotide binding and markedly slows down nucleotide release at NBD2. Login to comment
221 Evaluation of ADP release by wild-type and D793E, E1455D single and double mutations using 8-azido-[␣-32 P]ATP. Login to comment
234 The LTC4 labeling profile of the E1455D mutant in the absence of nucleotide was also similar to that observed for wild-type MRP1 (Fig. 6B, lanes 2 and 11). Login to comment
238 The double D793E/E1455D mutant behaved in a manner very similar to that of the D793E single mutation, with the exception that ATP␥S retained some ability to diminish LTC4 labeling (Fig. 6B, lanes 6-11). Login to comment
239 Thus these results together with those of vanadate trapping experiments are again consistent with the suggestion that increased trapping of ADP by the D793E mutant NBD1 decreases the nucleotide binding and hydrolysis by wild-type and E1455D mutant NBD2 and prevents conversion to a low affinity transition state. Login to comment
249 In contrast, the NBD2 mutations (E1455S, E1455N, E1455Q, and E1455L) like E1455D completely abolished LTC4 transport (Fig. 7B). Login to comment
255 Effect of ATP␥S or ADP trapping on [3 H]LTC4 photolabeling by wild-type MRP1, D793E, E1455D, and D793E/E1455D mutant proteins. Login to comment
256 Wild-type MRP1, D793E (A) and E1455D, D793E/ E1455D (B) membrane proteins (75 ␮g) were incubated in transport buffer containing 5 mM MgCl2 at 23 °C for 20 min in the absence (-) or presence (ϩ) of ATP (1 mM), vanadate (1 mM), or ATP␥S (4 mM), alone or in combination, prior to addition of [3 H]LTC4 (200 nM, 0.13 ␮Ci). Login to comment
263 The effect of the non-conservative mutations on nucleotide binding and ADP trapping was examined at 37 °C using 5 ␮M 8-azido-[␣-32 P]ATP in the presence or absence of vanadate, exactly as described for the D793E and E1455D mutations. Login to comment
268 Like the E1455D mutant, the E1455S, E1455N, and E1455L mutations resulted in strong vanadate-independent photolabeling of NBD2 and increased vanadate-dependent photolabeling of NBD1 (Fig. 8C, lanes 3, 4, and 7-10). Login to comment
296 The E1455D mutation essentially eliminated transport of LTC4 as observed previously with mutations that inactivate NBD2 (Fig. 2B). Login to comment
302 Thus the E1455D mutation substantially increased affinity for both ATP and ADP. Login to comment
312 The E1455D mutation also strongly enhanced nucleotide binding and vanadate-dependent trapping at the co-expressed wild-type NBD1 (Fig. 3, B and C). Login to comment
319 Contrary to expectation, the markedly increased 8-azido-[␣-32 P]ADP binding by the E1455D mutant NBD2 was accompanied by increased rather than decreased ADP photolabeling of the co-expressed wild-type NBD1, suggesting that both NBDs could be simultaneously occupied by ADP. Login to comment
322 We also found that the E1455D mutation drastically decreased nucleotide release at 37 °C, as evidenced by an inability to displace either prebound 8-azido-[␣- 32 P]ATP (Fig. 5) or 8-azido-[␣-32 P]ADP (data not shown) with a large excess of cold nucleotide. Login to comment
324 In the absence of nucleotides, LTC4 photolabeling was unaffected by the E1455D mutation. Login to comment
327 This is consistent with stable vanadate-independent binding of nucleotide by the E1455D mutant NBD2 (Fig. 3B). Login to comment
328 In addition, ATP␥S was much more effective at reducing binding by the E1455D mutant when compared with the wild-type protein. Login to comment
330 Thus the loss of LTC4 transport activity appears to be attributable to the impaired ability of the E1455D mutant to release nucleotide from NBD2 and to re-establish a high affinity binding state. Login to comment
331 Like the single mutations, the D793E/E1455D mutation had no effect on LTC4 binding in the absence of nucleotides. Login to comment
332 However, in the presence of ATP or ATP plus vanadate, the D793E mutation in the double mutant abrogated the shift from a high to a low affinity binding state, despite the potentiating effect observed with the E1455D single mutation. Login to comment
333 Paradoxically, a decrease of LTC4 binding by the D793E/E1455D double mutant protein was still observed in the presence of ATP␥S (Fig. 6B). Login to comment
342 Like the E1455D mutation, the substitution of Glu1455 with Gln, Asn, Leu, and Ser completely abolished LTC4 transport activity (Fig. 7B). Login to comment
343 However, in contrast to the E1455D mutation, which at 4 °C increased ATP binding at NBD2, binding was decreased by the Gln, Ser, and Leu mutations, whereas the Asn mutation had little or no effect relative to wild-type (Fig. 8A, lanes 4, 6, 8, and 9). Login to comment
345 At 37 °C, Ser, Asn, and Leu mutations of Glu1455 strongly increased nucleotide binding and trapping at both NBDs in the presence and absence of vanadate, as observed with the E1455D mutation, whereas binding by the Gln mutant protein, although increased relative to wild-type MRP1, remained strongly vanadate-dependent (Fig. 8C). Login to comment
346 This suggests that the Gln mutant, unlike the other mutant proteins including the E1455D mutant, is able to hydrolyze ATP and probably release ADP in the absence of vanadate. Login to comment

[hide] Functional interactions between nucleotide binding... Mol Pharmacol. 2005 Jun;67(6):1944-53. Epub 2005 Mar 8. Payen L, Gao M, Westlake C, Theis A, Cole SP, Deeley RG
Functional interactions between nucleotide binding domains and leukotriene C4 binding sites of multidrug resistance protein 1 (ABCC1).
Mol Pharmacol. 2005 Jun;67(6):1944-53. Epub 2005 Mar 8., [PMID:15755910]

Abstract [show]
Multidrug resistance protein 1 (MRP1) is a member of the "C" branch of the ATP-binding cassette transporter superfamily. The NH(2)-proximal nucleotide-binding domain (NBD1) of MRP1 differs functionally from its COOH-proximal domain (NBD2). NBD1 displays intrinsic high-affinity ATP binding and little ATPase activity. In contrast, ATP binding to NBD2 is strongly dependent on nucleotide binding by NBD1, and NBD2 is more hydrolytically active. We have demonstrated that occupancy of NBD2 by ATP or ADP markedly decreased substrate binding by MRP1. We have further explored the relationship between nucleotide and substrate binding by examining the effects of various ATP analogs and ADP trapping, as well as mutations in conserved functional elements in the NBDs, on the ability of MRP1 to bind the photoactivatable, high-affinity substrate cysteinyl leukotriene C(4) (LTC(4))(.) Overall, the results support a model in which occupancy of both NBD1 and NBD2 by ATP results in the formation of a low-affinity conformation of the protein. However, nonhydrolyzable ATP analogs (beta,gamma-imidoadenosine 5'-triphosphate and adenylylmethylene diphosphonate) failed to substitute for ATP or adenosine 5'-O-(thiotriphosphate) (ATPgammaS) in decreasing LTC(4) photolabeling. Furthermore, mutations of the signature sequence in either NBD that had no apparent effect on azido-ATP binding abrogated the formation of a low-affinity substrate binding state in the presence of ATP or ATPgammaS. We suggest that the effect of these mutations, and possibly the failure of some ATP analogs to decrease LTC(4) binding, may be attributable to an inability to elicit a conformational change in the NBDs that involves interactions between the signature sequence and the gamma-phosphate of the bound nucleotide.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
46 The reciprocal E1455D mutation of NBD2 increased the affinity of NBD2 for both azido-ATP and -ADP, resulting in prolonged binding of both nucleotides. Login to comment

[hide] Structure of the human multidrug resistance protei... J Mol Biol. 2006 Jun 16;359(4):940-9. Epub 2006 May 2. Ramaen O, Leulliot N, Sizun C, Ulryck N, Pamlard O, Lallemand JY, Tilbeurgh H, Jacquet E
Structure of the human multidrug resistance protein 1 nucleotide binding domain 1 bound to Mg2+/ATP reveals a non-productive catalytic site.
J Mol Biol. 2006 Jun 16;359(4):940-9. Epub 2006 May 2., 2006-06-16 [PMID:16697012]

Abstract [show]
Human multidrug resistance protein 1 (MRP1) is a membrane protein that belongs to the ATP-binding cassette (ABC) superfamily of transport proteins. MRP1 contributes to chemotherapy failure by exporting a wide range of anti-cancer drugs when over expressed in the plasma membrane of cells. Here, we report the first high-resolution crystal structure of human MRP1-NBD1. Drug efflux requires energy resulting from hydrolysis of ATP by nucleotide binding domains (NBDs). Contrary to the prokaryotic NBDs, the extremely low intrinsic ATPase activity of isolated MRP1-NBDs allowed us to obtain the structure of wild-type NBD1 in complex with Mg2+/ATP. The structure shows that MRP1-NBD1 adopts a canonical fold, but reveals an unexpected non-productive conformation of the catalytic site, providing an explanation for the low intrinsic ATPase activity of NBD1 and new hypotheses on the cooperativity of ATPase activity between NBD1 and NBD2 upon heterodimer formation.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
129 The role of the catalytic carboxylate was examined from NBD1- Asp793 and NBD2-Glu1455 mutations: the Asp793Glu mutation in NBD1 enhances its hydrolytic capacity, whereas the Glu1455Asp mutation in NBD2 results in an increased affinity of NBD2 for ATP, but a reduced hydrolytic activity.25 Our model shows that the orientation of His1486 could be sensitive to the conformation of His827, which is constrained by a strong hydrogen bond with Asp793, itself possibly stabilized by the participation of Asp792 to the nucleotide-binding site in the Mg2C /ATP-bound state. Login to comment

[hide] A molecular understanding of ATP-dependent solute ... Cancer Metastasis Rev. 2007 Mar;26(1):15-37. Chang XB
A molecular understanding of ATP-dependent solute transport by multidrug resistance-associated protein MRP1.
Cancer Metastasis Rev. 2007 Mar;26(1):15-37., [PMID:17295059]

Abstract [show]
Over a million new cases of cancers are diagnosed each year in the United States and over half of these patients die from these devastating diseases. Thus, cancers cause a major public health problem in the United States and worldwide. Chemotherapy remains the principal mode to treat many metastatic cancers. However, occurrence of cellular multidrug resistance (MDR) prevents efficient killing of cancer cells, leading to chemotherapeutic treatment failure. Numerous mechanisms of MDR exist in cancer cells, such as intrinsic or acquired MDR. Overexpression of ATP-binding cassette (ABC) drug transporters, such as P-glycoprotein (P-gp or ABCB1), breast cancer resistance protein (BCRP or ABCG2) and/or multidrug resistance-associated protein (MRP1 or ABCC1), confers an acquired MDR due to their capabilities of transporting a broad range of chemically diverse anticancer drugs. In addition to their roles in MDR, there is substantial evidence suggesting that these drug transporters have functions in tissue defense. Basically, these drug transporters are expressed in tissues important for absorption, such as in lung and gut, and for metabolism and elimination, such as in liver and kidney. In addition, these drug transporters play an important role in maintaining the barrier function of many tissues including blood-brain barrier, blood-cerebral spinal fluid barrier, blood-testis barrier and the maternal-fetal barrier. Thus, these ATP-dependent drug transporters play an important role in the absorption, disposition and elimination of the structurally diverse array of the endobiotics and xenobiotics. In this review, the molecular mechanism of ATP-dependent solute transport by MRP1 will be addressed.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
242 In contrast, mutation of the putative catalytic residue E1455 to a short chain D residue, E1455D, markedly increased the affinity of the mutated NBD2 for ATP while decreasing its ability to hydrolyze ATP [62], leading to significantly increased α-32 P-ATP labeling regardless of whether Vi is present or not [62]. Login to comment
243 Binding of ATP, ATP + Vi, or ATPγS to E1455D significantly inhibited the LTC4 labeling [62], further supporting the above hypothesis that occupancy of both NBD1 and NBD2 by nucleotide binding without hydrolysis may be sufficient to transport the bound substrate across membrane bilayer. Login to comment
245 For example, although ATPγS binding to wild-type or E1455D-mutated MRP1 significantly inhibited LTC4 labeling [62], ATPγS itself did not support the ATP-dependent LTC4 or E217βG transport [33, 47]; ATP can efficiently bind to E1455D, D793E/ E1455D or E1455L [62, 144], but mutation of this putative catalytic residue abolished the ATP-dependent solute transport [62, 144]. Login to comment
261 This conclusion is further supported by mutation of the putative catalytic residue E1455 in NBD2 that all the mutants, including E1455S, E1455Q, E1455N, E1455L and E1455D, lost their abilities to transport LTC4 across membrane bilayer [62, 144]. Login to comment

[hide] Molecular mechanism of ATP-dependent solute transp... Methods Mol Biol. 2010;596:223-49. Chang XB
Molecular mechanism of ATP-dependent solute transport by multidrug resistance-associated protein 1.
Methods Mol Biol. 2010;596:223-49., [PMID:19949927]

Abstract [show]
Millions of new cancer patients are diagnosed each year and over half of these patients die from this devastating disease. Thus, cancer causes a major public health problem worldwide. Chemotherapy remains the principal mode to treat many metastatic cancers. However, occurrence of cellular multidrug resistance (MDR) prevents efficient killing of cancer cells, leading to chemotherapeutic treatment failure. Over-expression of ATP-binding cassette transporters, such as P-glycoprotein, breast cancer resistance protein and/or multidrug resistance-associated protein 1 (MRP1), confers an acquired MDR due to their capabilities of transporting a broad range of chemically diverse anticancer drugs across the cell membrane barrier. In this review, the molecular mechanism of ATP-dependent solute transport by MRP1 will be addressed.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
151 However, binding of poorly hydrolysable ATP analog ATPgS to wt MRP1 significantly inhibits the 3 H-LTC4 labeling (99, 100), implying that ATPgS binding might be sufficient to transport the bound LTC4 from high to low affinity site. This conclusion was further supported by the fact that ATPgS or ATP binding to the incompetent E1455D or E1455Q mutants, which were unable to hydrolyze the bound ATP, significantly inhibited the 3 H-LTC4 labeling (100, 156, 157). Login to comment
158 Conversely, mutation of the putative catalytic residue E1455 to a short chain D residue, E1455D, markedly increased the affinity of the mutated NBD2 for ATP while decreased its ability to hydrolyze ATP (100), leading to significantly increase the a-32 P-ATP labeling regardless of whether Vi is present or not (100). Login to comment
159 Binding of ATP, ATP + Vi, or ATPgS to E1455D significantly inhibited the LTC4 labeling (100), further supporting the above hypothesis that occupancies of both NBD1 and NBD2 by nucleotide binding without hydrolysis may be sufficient to transport the bound substrate across membrane bilayer. Login to comment

[hide] Structural and functional properties of human mult... Curr Med Chem. 2011;18(3):439-81. He SM, Li R, Kanwar JR, Zhou SF
Structural and functional properties of human multidrug resistance protein 1 (MRP1/ABCC1).
Curr Med Chem. 2011;18(3):439-81., [PMID:21143116]

Abstract [show]
Multidrug ABC transporters such as P-glycoprotein (P-gp/MDR1/ABCB1) and multidrug resistance protein 1 (MRP1/ABCC1) play an important role in the extrusion of drugs from the cell and their overexpression can be a cause of failure of anticancer and antimicrobial chemotherapy. Recently, the mouse P-gp/Abcb1a structure has been determined and this has significantly enhanced our understanding of the structure-activity relationship (SAR) of mammalian ABC transporters. This paper highlights our current knowledge on the structural and functional properties and the SAR of human MRP1/ABCC1. Although the crystal structure of MRP1/ABCC1 has yet to be resolved, the current topological model of MRP1/ABCC1 contains two transmembrane domains (TMD1 and TMD2) each followed by a nucleotide binding domain (NBD) plus a third NH2-terminal TMD0. MRP1/ABCC1 is expressed in the liver, kidney, intestine, brain and other tissues. MRP1/ABCC1 transports a structurally diverse array of important endogenous substances (e.g. leukotrienes and estrogen conjugates) and xenobiotics and their metabolites, including various conjugates, anticancer drugs, heavy metals, organic anions and lipids. Cells that highly express MRP1/ABCC1 confer resistance to a variety of natural product anticancer drugs such as vinca alkaloids (e.g. vincristine), anthracyclines (e.g. etoposide) and epipodophyllotoxins (e.g. doxorubicin and mitoxantrone). MRP1/ABCC1 is associated with tumor resistance which is often caused by an increased efflux and decreased intracellular accumulation of natural product anticancer drugs and other anticancer agents. However, most compounds that efficiently reverse P-gp/ABCB1-mediated multidrug resistance have only low affinity for MRP1/ABCC1 and there are only a few effective and relatively specific MRP1/ABCC1 inhibitors available. A number of site-directed mutagenesis studies, biophysical and photolabeling studies, SAR and QSAR, molecular docking and homology modeling studies have documented the role of multiple residues in determining the substrate specificity and inhibitor selectivity of MRP1/ABCC1. Most of these residues are located in the TMs of TMD1 and TMD2, in particular TMs 4, 6, 7, 8, 10, 11, 14, 16, and 17, or in close proximity to the membrane/cytosol interface of MRP1/ABCC1. The exact transporting mechanism of MRP1/ABCC1 is unclear. MRP1/ABCC1 and other multidrug transporters are front-line mediators of drug resistance in cancers and represent important therapeutic targets in future chemotherapy. The crystal structure of human MRP1/ABCC1 is expected to be resolved in the near future and this will provide an insight into the SAR of MRP1/ABCC1 and allow for rational design of anticancer drugs and potent and selective MRP1/ABCC1 inhibitors.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
654 The Asp793Glu mutation in NBD1 enhanced its hydrolytic capacity, whereas the Glu1455Asp mutation in NBD2 resulted in an increased affinity of NBD2 for ATP, but a decreased hydrolytic activity [322]. Login to comment
673 The Glu1455Asp mutant with a reciprocal mutation of NBD2 exhibited an increased affinity for both azido-ATP and -ADP. Login to comment