ABCC1 p.Ser604Thr
| Predicted by SNAP2: | A: D (75%), C: D (85%), D: D (95%), E: D (95%), F: D (95%), G: D (85%), H: D (91%), I: D (95%), K: D (95%), L: D (95%), M: D (91%), N: D (91%), P: D (95%), Q: D (91%), R: D (95%), T: D (85%), V: D (91%), W: D (95%), Y: D (95%), |
| Predicted by PROVEAN: | A: N, C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: N, P: D, Q: D, R: D, T: N, V: D, W: D, Y: D, |
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[hide] Transmembrane helix 11 of multidrug resistance pro... Biochemistry. 2004 Jul 27;43(29):9413-25. Zhang DW, Nunoya K, Vasa M, Gu HM, Theis A, Cole SP, Deeley RG
Transmembrane helix 11 of multidrug resistance protein 1 (MRP1/ABCC1): identification of polar amino acids important for substrate specificity and binding of ATP at nucleotide binding domain 1.
Biochemistry. 2004 Jul 27;43(29):9413-25., 2004-07-27 [PMID:15260484]
Abstract [show]
Human multidrug resistance protein 1 (MRP1) is an ATP binding cassette (ABC) transporter that confers resistance to many natural product chemotherapeutic agents and can transport structurally diverse conjugated organic anions. MRP1 has three polytopic transmembrane domains (TMDs) and a total of 17 TM helices. Photolabeling and mutagenesis studies of MRP1 indicate that TM11, the last helix in the second TMD, may form part of the protein's substrate binding pocket. We have demonstrated that certain polar residues within a number of TM helices, including Arg(593) in TM11, are determinants of MRP1 substrate specificity or overall activity. We have now extended these analyses to assess the functional consequences of mutating the remaining seven polar residues within and near TM11. Mutations Q580A, T581A, and S585A in the predicted outer leaflet region of the helix had no detectable effect on function, while mutation of three residues close to the membrane/cytoplasm interface altered substrate specificity. Two of these mutations affected only drug resistance. N597A increased and decreased resistance to vincristine and VP-16, respectively, while S605A decreased resistance to vincristine, VP-16 and doxorubicin. The third, S604A, selectively increased 17beta-estradiol 17-(beta-d-glucuronide) (E(2)17betaG) transport. In contrast, elimination of the polar character of the residue at position 590 (Asn in the wild-type protein) uniformly impaired the ability of MRP1 to transport potential physiological substrates and to confer resistance to three different classes of natural product drugs. Kinetic and photolabeling studies revealed that mutation N590A not only decreased the affinity of MRP1 for cysteinyl leukotriene 4 (LTC(4)) but also substantially reduced the binding of ATP to nucleotide binding domain 1 (NBD1). Thus, polar interactions involving residues in TM11 influence not only the substrate specificity of MRP1 but also an early step in the proposed catalytic cycle of the protein.
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No. Sentence Comment
55 Mutations Q580A, T581A, S604T, and S605A were generated using the Transformer- Site-Directed Mutagenesis kit (CLONTECH, Palo Alto, CA).
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ABCC1 p.Ser604Thr 15260484:55:24
status: NEW59 They are as follows: Q580A (5'-C ATC CTG GAT GCC GCG ACG GCC TTC GTG TC-3'), T581A (5'-CTG GAT GCC CAG GCG GCC TTC GTG TCT TTG-3'), S605A (5'-C ATG GTC ATC AGC GCG ATC GTG CAG GCG-3'), and S604T (5'-CCC ATG GTC ATC ACT AGT ATC GTG CAG GCG-3').
X
ABCC1 p.Ser604Thr 15260484:59:189
status: NEW158 Effect of Mutations S604T, N590Q, and N590D on Transport of [3 H]LTC4 and [3 H]E217 G by Wild-Type MRP1.
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ABCC1 p.Ser604Thr 15260484:158:20
status: NEW161 Ser604 was mutated to Thr, and Asn590 was converted to Asp and Gln.
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ABCC1 p.Ser604Thr 15260484:161:0
status: NEW165 Mutations N590Q, N590D, and S604T had no effect on transport of the two substrates.
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ABCC1 p.Ser604Thr 15260484:165:28
status: NEW203 Mutations Q580A, T581A, S585A, S604A, and S604T had no significant effect on the ability of MRP1 to confer resistance to any drug tested.
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ABCC1 p.Ser604Thr 15260484:203:42
status: NEW270 In addition, although mutation S604T had no effect on any of the MRP1 functions tested, converting Ser604 to Ala decreased the apparent Km values and increased Vmax for E217 G uptake.
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ABCC1 p.Ser604Thr 15260484:270:31
status: NEW