ABCC1 p.Leu1430Arg
| Predicted by SNAP2: | A: D (80%), C: D (63%), D: D (95%), E: D (91%), F: D (66%), G: D (95%), H: D (91%), I: D (63%), K: D (95%), M: D (63%), N: D (91%), P: D (95%), Q: D (91%), R: D (95%), S: D (91%), T: D (91%), V: D (71%), W: D (91%), Y: D (91%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: D, F: D, G: D, H: D, I: N, K: D, M: N, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D, |
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[hide] Functional studies on the MRP1 multidrug transport... Anticancer Res. 2004 Mar-Apr;24(2A):449-55. Szentpetery Z, Sarkadi B, Bakos E, Varadi A
Functional studies on the MRP1 multidrug transporter: characterization of ABC-signature mutant variants.
Anticancer Res. 2004 Mar-Apr;24(2A):449-55., [PMID:15152943]
Abstract [show]
BACKGROUND: MRP1 is a key multidrug resistance ATP-binding Cassette (ABC) transporter in tumor cells. A functionally important signature motif is conserved within all ABC domains. Our current studies aimed to elucidate the role of these motifs in the cooperation of MRP1 ABC domains. MATERIALS AND METHODS: We designed human MRP1 mutants based on a bacterial ABC structure. Conserved leucines (Leu) were replaced by arginines (Arg), while glycines (Gly) were substituted for aspartic acids (Asp). The activity of these mutants was assayed by measuring ATPase activity and vesicular transport. ATP-binding and transition-state formation were studied by a photoreactive ATP analog. RESULTS: The Leu to Arg mutants retained both ATPase and transport activity, while the Gly to Asp mutants were inactive in all functional assays, while showing normal ATP-binding. CONCLUSION: Our results reinforce the notion that a single mutation in one of the ABC-signature regions affects the function of the whole protein. The relative role of the conservative leucines and glycines in MRP1 indicates a similar three-dimensional structure within the catalytic center of various ABC proteins.
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No. Sentence Comment
46 The conserved leucines of the LSGGQ motifs were replaced by arginines (L768R, L1430R) and the conserved glycines in the fourth position of the signature motifs were substituted for aspartic acids (G771D, G1433D).
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ABCC1 p.Leu1430Arg 15152943:46:78
status: NEW122 In Figure 2 we document that, when measuring the ATPase activity of the L768R and L1430R signature mutants, we found that they possessed a significant level of vanadate-sensitive, drug-stimulated ATPase activity.
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ABCC1 p.Leu1430Arg 15152943:122:82
status: NEW128 As shown in Figure 3, in harmony with the ATPase activity measurements, the L768R and L1430R signature mutants presented 87-89 % of the transport activity of the wild-type MRP1, while the G771D and G1433D signature mutants had only a negligible level of ATP-dependent LTC4 uptake, similar to that found in the ‚-galactosidase- expressing control membranes.
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ABCC1 p.Leu1430Arg 15152943:128:86
status: NEW129 All these functional studies indicated that the catalytic cycles of the L768R and L1430R mutants were similar to the wild-type, while that of the G771D and G1433D mutants were seriously diminished.
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ABCC1 p.Leu1430Arg 15152943:129:82
status: NEW172 We chose leucine, a residue not oriented towards the ATP, which we substituted for arginine (L768R, L1430R).
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ABCC1 p.Leu1430Arg 15152943:172:100
status: NEW178 We found that the ATPase and transport activity of the L768R and L1430R signature mutants, reflecting the whole catalytic cycle of MRP1, were not significantly different from that of the wild-type MRP1.
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ABCC1 p.Leu1430Arg 15152943:178:65
status: NEW[hide] The role of the conserved glycines of ATP-binding ... J Biol Chem. 2004 Oct 1;279(40):41670-8. Epub 2004 Jul 12. Szentpetery Z, Kern A, Liliom K, Sarkadi B, Varadi A, Bakos E
The role of the conserved glycines of ATP-binding cassette signature motifs of MRP1 in the communication between the substrate-binding site and the catalytic centers.
J Biol Chem. 2004 Oct 1;279(40):41670-8. Epub 2004 Jul 12., 2004-10-01 [PMID:15252017]
Abstract [show]
A key element of the structural model of ABC-ATP-ases is the interaction of the two ABC domains. They complement each other's active sites in a way that the ABC signature motif (LSGGQ) of one subunit interacts with the gamma-phosphate of the ATP, bound at the Walker motifs of the opposite subunit. In the present study, the conserved glycines in the fourth position of the LSGGQ motifs of human MRP1 were substituted for aspartic acids (G771D and G1433D), the mutants were expressed in Sf9 insect cells, and the nucleotideas well as the transported substrate-protein interactions were studied. We found that these transport- and ATPase-incompetent mutants showed no nucleotide trapping under any of the conditions examined. However, when measuring the effect of nucleotide and transported substrates on the vanadate-induced cleavage reactions, we found that the effect of substrates on the cleavage reactions was significantly different in the mutant MRP1 proteins than in the wild type. Although the transported substrates (e.g. etoposide + oxidized glutathione) stimulated the formation of the posthydrolytic complex in the wild type, this reaction was inhibited in the signature mutants. Our study also revealed that a similar mutation in the ABC signature of either ABC unit resulted in the same effect. We suggest that the conserved glycine residues in both LSGGQ segments are part of the conformational network, which is responsible for the accelerated hydrolytic activity upon interaction of the protein with its transported substrates. This intramolecular communication between the substrate-binding site and the catalytic centers is assumed to be a general feature of the molecular mechanism of ABC transporters.
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No. Sentence Comment
184 These mutations affect the conserved LSGGQ motifs in either ABC domain; the leucines were replaced with arginines (L768R and L1430R), and the glycines in position 4 were substituted with glutamic acids (G771D and G1433D).
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ABCC1 p.Leu1430Arg 15252017:184:125
status: NEW[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.
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No. Sentence Comment
181 Mutation of the first residue in LSGGQ signature sequence to an R residue, L768R in NBD1 or L1430R in NBD2, retained both ATPase activity and ATP-dependent solute transport activity [116], indicating that this L residue is not crucial for the NBD1·ATP2·NBD2 sandwich structure formation.
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ABCC1 p.Leu1430Arg 17295059:181:92
status: NEW