ABCB4 p.Glu1207Gln
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PMID: 23593265
[PubMed]
Ellinger P et al: "Detergent screening and purification of the human liver ABC transporters BSEP (ABCB11) and MDR3 (ABCB4) expressed in the yeast Pichia pastoris."
No.
Sentence
Comment
106
Oligonucleotide Sequence 59R 39 pSGP18-2m-ori-S1 TAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTAAATATTGCGAATACCGCTTCCACAAACATTG pSGP18-2m-ori-S2 AACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGTTATTTCACACCGCATATATCGGATCGTACT BSEP-HR-PP-S1 ATCAAAAAACAACTAATTATTCGAACGAGGTAAAAGAATGTCTGACTCAGTAATTCTTCGAAGT ATA BSEP-HR-PP-S2 ACGTTTGGACCTTGGAAAAGACTTCTAAGGAGTTGGAGGCACTGATGGGGGATCCAGTGGTGACTAGTTT MDR3-HR-PP-S1 ATCAAAAAACAACTAATTATTCGAACGAGGTAAAAGAATGGATCTTGAGGCGGCAAAGAACGGAACA MDR3-HR-PP-S2 ACGTTTGGACCTTGGAATAAGACTTCTAAGGAGTTGGAGGCTAAGTTCTGTGTTCCAGCCTGGACACTGACCATTGAAAAATAG YEpN14HIS-BSEP-S2 GAATAAGGTAAACATGGTAGCGATGTCGACCTCGAGACGCGTCTAACTGATGGGGGATCCAGTGGTGACT YEpN14HIS-MDR3-S2 GAATAAGGTAAACATGGTAGCGATGTCGACCTCGAGACGCGTCTATAAGTTCTGTGTCCCAGCCTGGACACTGACCATT GFP-BSEP-HR-S1 AGCCTACTACAAACTAGTCACCACTGGATCCCCCATCAGTGGTGGTGGTCGACGGATCCCCGGGTTA GFP-PP-HR-S2 ACGTTTGGACCTTGGAATAAGACTTCTAAGGAGTTGGAGGCTATTATTTGTATAGTTCATCCATGCCATGT GFP-MDR3-HR-S1 TTTCAATGGTCAGTGTCCAGGCTGGAACAAAGAGACAAGGTGGTGGTCGACGGATCCCCGGGTTA MDR3-E558Q S1 GATCCTTCTGCTGGATCAAGCCACGTCAGCATTGGACAC MDR3-E558Q S2 GTGTCCAATGCTGACGTGGCTTGATCCAGCAGAAGGATC MDR3-E1207Q S1 CAAATCCTCCTGTTGGATCAAGCTACATCAGCTCTGGATAC MDR3-E1207Q S2 GTATCCAGAGCTGATGTAGCTTGATCCAACAGGAGGATTTG doi:10.1371/journal.pone.0060620.t001 Preparation of crude membrane vesicles for protein purification 100 g batches of P. pastoris cells expressing BSEP or MDR3 were thawed on ice, washed with ddH2O and re-suspended at a concentration of 0.5 g cells/ml in homogenization buffer containing protease inhibitor cocktail (Roche).
X
ABCB4 p.Glu1207Gln 23593265:106:1128
status: NEWX
ABCB4 p.Glu1207Gln 23593265:106:1185
status: NEW248 The ATPase inactive mutant (E558Q, E1207Q, further called EQ/EQ mutant) exhibited basal ATPase activity comparable to the wild-type protein.
X
ABCB4 p.Glu1207Gln 23593265:248:35
status: NEW280 Molecular weight markers are shown on the left. B Normalized ATPase activity of MDR3 wild-type (black) and of an ATPase deficient mutant (E558Q E1207Q, white) in FC-16 without and with different phospholipids.
X
ABCB4 p.Glu1207Gln 23593265:280:144
status: NEW322 This observation was sustained by analysis of an ATP hydrolysis deficient EQ double mutant (E558Q, E1207Q).
X
ABCB4 p.Glu1207Gln 23593265:322:99
status: NEW105 Oligonucleotide Sequence 59R 39 pSGP18-2m-ori-S1 TAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTAAATATTGCGAATACCGCTTCCACAAACATTG pSGP18-2m-ori-S2 AACGCGGCCTTTTTACGGTTCCTGGCCTTTTGCTGGCCTTTTGCTCACATGTTATTTCACACCGCATATATCGGATCGTACT BSEP-HR-PP-S1 ATCAAAAAACAACTAATTATTCGAACGAGGTAAAAGAATGTCTGACTCAGTAATTCTTCGAAGT ATA BSEP-HR-PP-S2 ACGTTTGGACCTTGGAAAAGACTTCTAAGGAGTTGGAGGCACTGATGGGGGATCCAGTGGTGACTAGTTT MDR3-HR-PP-S1 ATCAAAAAACAACTAATTATTCGAACGAGGTAAAAGAATGGATCTTGAGGCGGCAAAGAACGGAACA MDR3-HR-PP-S2 ACGTTTGGACCTTGGAATAAGACTTCTAAGGAGTTGGAGGCTAAGTTCTGTGTTCCAGCCTGGACACTGACCATTGAAAAATAG YEpN14HIS-BSEP-S2 GAATAAGGTAAACATGGTAGCGATGTCGACCTCGAGACGCGTCTAACTGATGGGGGATCCAGTGGTGACT YEpN14HIS-MDR3-S2 GAATAAGGTAAACATGGTAGCGATGTCGACCTCGAGACGCGTCTATAAGTTCTGTGTCCCAGCCTGGACACTGACCATT GFP-BSEP-HR-S1 AGCCTACTACAAACTAGTCACCACTGGATCCCCCATCAGTGGTGGTGGTCGACGGATCCCCGGGTTA GFP-PP-HR-S2 ACGTTTGGACCTTGGAATAAGACTTCTAAGGAGTTGGAGGCTATTATTTGTATAGTTCATCCATGCCATGT GFP-MDR3-HR-S1 TTTCAATGGTCAGTGTCCAGGCTGGAACAAAGAGACAAGGTGGTGGTCGACGGATCCCCGGGTTA MDR3-E558Q S1 GATCCTTCTGCTGGATCAAGCCACGTCAGCATTGGACAC MDR3-E558Q S2 GTGTCCAATGCTGACGTGGCTTGATCCAGCAGAAGGATC MDR3-E1207Q S1 CAAATCCTCCTGTTGGATCAAGCTACATCAGCTCTGGATAC MDR3-E1207Q S2 GTATCCAGAGCTGATGTAGCTTGATCCAACAGGAGGATTTG doi:10.1371/journal.pone.0060620.t001 Preparation of crude membrane vesicles for protein purification 100 g batches of P. pastoris cells expressing BSEP or MDR3 were thawed on ice, washed with ddH2O and re-suspended at a concentration of 0.5 g cells/ml in homogenization buffer containing protease inhibitor cocktail (Roche).
X
ABCB4 p.Glu1207Gln 23593265:105:1128
status: NEWX
ABCB4 p.Glu1207Gln 23593265:105:1185
status: NEW247 The ATPase inactive mutant (E558Q, E1207Q, further called EQ/EQ mutant) exhibited basal ATPase activity comparable to the wild-type protein.
X
ABCB4 p.Glu1207Gln 23593265:247:35
status: NEW279 Molecular weight markers are shown on the left. B Normalized ATPase activity of MDR3 wild-type (black) and of an ATPase deficient mutant (E558Q E1207Q, white) in FC-16 without and with different phospholipids.
X
ABCB4 p.Glu1207Gln 23593265:279:144
status: NEW321 This observation was sustained by analysis of an ATP hydrolysis deficient EQ double mutant (E558Q, E1207Q).
X
ABCB4 p.Glu1207Gln 23593265:321:99
status: NEW
PMID: 25533467
[PubMed]
Kluth M et al: "A mutation within the extended X loop abolished substrate-induced ATPase activity of the human liver ATP-binding cassette (ABC) transporter MDR3."
No.
Sentence
Comment
118
RESULTS Expression and Purification of the Human ABC Transporter MDR3 in P. pastoris-Previously, we described the expression of wild type MDR3 and the ATP hydrolysis-deficient mutant (E558Q/E1207Q, later called the EQ/EQ mutant) in the methylotrophic yeast P. pastoris.
X
ABCB4 p.Glu1207Gln 25533467:118:190
status: NEW156 A, human wild type MDR3, the E558Q/E1207Q double mutant, and the Q1174E mutant purified from P. pastoris.
X
ABCB4 p.Glu1207Gln 25533467:156:35
status: NEW182 of purifications Km (MgATP) vmax kcat mg/100 g wet cell weight mM nmol minafa;1 mgafa;1 safa;1 Wild type 6.3 afe; 1.2 8 2.17 afe; 0.20 354 afe; 13 0.83 afe; 0.03 1.78 afe; 0.10a 536 afe; 11a 1.26 afe; 0.03a Wild type-BODIPY 1.26 afe; 0.10 186 afe; 6 0.44 afe; 0.01 1.43 afe; 0.19a 175 afe; 10a 0.41 afe; 0.02a E558Q/E1207Q 3.4 afe; 0.6 5 b b b Q1174E 2.0 afe; 0.2 5 1.04 afe; 0.15 286 afe; 16 0.64 afe; 0.04 Q1174E-BODIPY 0.74 afe; 0.10 198 afe; 5 0.46 afe; 0.01 a ATPase activity was in the presence of 300 òe;M DOPC lipids.
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ABCB4 p.Glu1207Gln 25533467:182:364
status: NEW270 ATPase Activity of Human MDR3 4904 JOURNAL OF BIOLOGICAL CHEMISTRY VOLUME 290ߦNUMBER 8ߦFEBRUARY 20, 2015 at SEMMELWEIS UNIV OF MEDICINE on December , Cross-linking of wild type MDR3 with maleimide-BODIPY blocks basal and PC-induced ATPase activity as demonstrated for MDR1 (33-35), whereas the ATP hydrolysis-deficient EQ double mutant (E558Q/E1207Q) showed no PC stimulation, and ATPase activity of the labeled EQ/EQ mutant was only marginally reduced (Table 1 and Fig. 1).
X
ABCB4 p.Glu1207Gln 25533467:270:356
status: NEW269 ATPase Activity of Human MDR3 4904 JOURNAL OF BIOLOGICAL CHEMISTRY VOLUME 290ߦNUMBER 8ߦFEBRUARY 20, 2015 at SEMMELWEIS UNIV OF MEDICINE on December , Cross-linking of wild type MDR3 with maleimide-BODIPY blocks basal and PC-induced ATPase activity as demonstrated for MDR1 (33-35), whereas the ATP hydrolysis-deficient EQ double mutant (E558Q/E1207Q) showed no PC stimulation, and ATPase activity of the labeled EQ/EQ mutant was only marginally reduced (Table 1 and Fig. 1).
X
ABCB4 p.Glu1207Gln 25533467:269:356
status: NEW