ABCMdb

ABCC1 p.Arg1166Ala

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Publications

PMID: 19015228 [PubMed] Conseil G et al: "Multiple roles of charged amino acids in cytoplasmic loop 7 for expression and function of the multidrug and organic anion transporter MRP1 (ABCC1)."

No. Sentence Comment

6 The properties of R1173A and E1184A were comparable with those of wild-type MRP1, whereas the remaining mutants were either poorly expressed (R1166A, D1183A) or exhibited reduced transport of one or more organic anions (E1144A, D1179A, K1181A, 1169 AAQA). Login to comment
59 Mutations were first generated in the pGEM-3Z-XmaI/MRP1 plasmid according to the manufacturer`s instructions with the following mutagenic primers (Integrated DNA Technologies, Inc., Coralville, IA), which also introduced an additional restriction site (substituted nucleotides are underlined): E1144A (5Ј-G AAG CGC CTC GCG TCG GTC AGC-3Ј); R1166A (5Ј C AGC GTC ATT GCA GCA TTC GAG GAG CAG-3Ј); R1166K (5Ј C AGC GTC ATT AAG GCC TTC GAG G-3Ј); 1169 EEQE-1169 AAQA (5Ј-C ATT CGA GCC TTC GCG GCA CAG GCA CGC TTC ATC C-3Ј); R1173A (5Ј-C GAG GAG CAG GAG GCA TTC ATC CAC CAG AG-3Ј); D1179A (5Ј-C CAC CAG AGT GCC CTT AAG GTG GAC G-3Ј), K1181A (5Ј-G AGT GAC CTG GCA GTC GAC GAG AAC C-3Ј); D1183A (5Ј-CTG AAG GTG GCC GAG AAC CAG-3Ј); D1183E (5Ј- CTG AAG GTG GAA GAG AAC CAG-3Ј); and E1184A (5Ј-GT GAC CTG AAG GTA GAC GCG AAC CAG AAG GCC-3Ј). Login to comment
120 These experiments showed that two of the Ala-substituted mutants (R1166A and D1183A) were consistently expressed at very low levels (10-20% of wild-type MRP1) (Fig. 2A). Login to comment
124 Confocal microscopy of intact HEK cells using the MRP1-specific mAb QCRL-3 showed that despite their low expression levels, at least a portion of the R1166A and D1183A mutant proteins were correctly routed to the plasma mem- brane (not shown). Login to comment
126 However, even at the lower temperature, expression of the R1166A and D1183A mutants remained substantially below that of wild-type MRP1; furthermore, plasma membrane routing of the mutants as well as wild-type MRP1 was impaired (data not shown). Login to comment
138 A, immunoblot of whole cell lysates (10 ␮g of protein) prepared from HEK 293T cells transfected with wild-type (WT-MRP1), and R1166A and D1183A mutant MRP1 expression vectors. Login to comment
140 MRP1 levels in whole-cell lysates were detected with mAb QCRL-1, and the relative protein expression levels are shown under the blot and were estimated by densitometry as described under Materials and Methods. B, immunoblots of whole-cell lysates (10 ␮g of protein) prepared from HEK 293T cells transfected with R1166A/K and D1183A/E mutant and wild-type (WT) MRP1 cDNA expression vectors. MRP1 proteins were detected with mAb QCRL-1 and lysates of untransfected cells were included as a negative control as above. Login to comment
192 The remaining five single mutants were either poorly expressed (R1166A, D1183A) (Fig. 2) or exhibited significantly reduced transport levels of one or more organic anions (E1144A, D1179A, K1181A) (Table 1). Login to comment
194 The poor expression of the R1166A and D1183A mutants indicates that Arg1166 and Asp1183 contribute to the stability of MRP1, probably by influencing the proper folding of the transporter during its biosynthesis. Login to comment
199 Elucidation of the post-translational events leading to poor expression of R1166A and D1183A and other low or nonexpressing MRP1 mutants is currently under investigation. Login to comment
238 Fanen et al. (1997) further found that transfected cells expressing CFTR-R1066C did not respond to cAMP stimulation, and, similar to what we observed with MRP1-R1166A, the mutant CFTR protein was poorly expressed. Login to comment
240 Likewise, we found that expression of MRP1-R1166A could not be improved under comparable conditions. Login to comment
244 Unlike the R1166A and D1183A mutants, the second group of functionally altered single Ala-substituted mutants (E1144A, D1179A, K1181A) and the 1169 AAQA triple mutant were all expressed at levels comparable with or greater than that of wild-type MRP1 (Fig. 4). Login to comment

PMID: 21143116 [PubMed] He SM et al: "Structural and functional properties of human multidrug resistance protein 1 (MRP1/ABCC1)."

No. Sentence Comment

726 These include Cys43 in TM1 [171], Thr73 in CL1 [366], Trp222 in L0 [279], Trp223 in L0 [279], Arg230 in L0 [279], Trp261 in L0 [271, 302, 338], Lys267 in L0 [271, 302, 338], Lys319 in TM6 [339], Tyr324 in TM6 [301], Lys332 in TM6 [166, 339-341], His335 in TM6, Asp336 in TM6 [339], Lys347 in TM6 [339], Lys396 in TM7 [339], Arg433 in TM8 [363], Asp436 in TM8 [339], Trp445 in TM8, Trp459 in TM8, Pro478 in TM9, Thr550 in TM10 [343], Trp553 in TM10 [344], Thr556 in TM10 [343], Pro557 in TM10 [345], Tyr568 in TM10 [343], Arg593 in TM11 [339], Phe594 in TM11 [300], Asn597 in TM11, Ser604 in TM11, Ser605 in TM11, Trp653 in NBD1 [351], Lys684 in Walker A motif of NBD1 [350, 364], Ser685 in Walker A motif of NBD1 [353], Arg723 in NBD1 [366], Gly771 in the ABC signature (C motif) of NBD1 [364], Asp792 in Walker B motif of NBD1 [361], Asp793 in Walker B motif of NBD1 [353], Ala989 in TM12 [367], Pro1120 in TM13, Pro1121 in TM13, Arg1058 at TM13/CL6 interface [366], Glu1089 in TM14, Lys1092 in TM14, Ser1097 in TM14, Asn1100 in TM14, Arg1138 in TM15 [354], Lys1141 in CL7 [354], Arg1142 in CL7 [354], Glu1144Ala in CL7 [355], Pro1150 in CL7 [345, 347, 348], Arg1166Ala in CL7 [355], Asp1179Ala in CL7 [355], Lys1181Ala in CL7 [355], Asp1183 in CL7 [355], Tyr1189 in CL7 [368], Tyr1190 in CL7 [368], Arg1197 in TM16 [357], Trp1198 in TM16 [344], Arg1202 in TM16 [357], Glu1204 in TM16 [357], Tyr1236 in TM17, Thr1241 in TM17, Thr1242 in TM17, Tyr1243 in TM17, Asn1245 in TM17, Trp1246 in TM17 [166, 339-341], Arg1249 in TM17 [342], Tyr1302 in NBD2 [351], Lys1333 in Walker A motif of NBD2 [350], Gly1433 in the ABC signature motif of NBD2 [352, 364], Glu1455 in Walker B motif of NBD2 [322], and His1486 in NBD2 [349]. Login to comment
809 The Arg1166Ala and Asp1183Ala/Glu mutants were poorly expressed, probably by affecting the proper folding of the protein during its biosynthesis. Login to comment