ABCMdb

ABCB6 p.Cys50Ala

None has been submitted yet.

Publications

PMID: 21199866 [PubMed] Fukuda Y et al: "Conserved intramolecular disulfide bond is critical to trafficking and fate of ATP-binding cassette (ABC) transporters ABCB6 and sulfonylurea receptor 1 (SUR1)/ABCC8."

No. Sentence Comment

61 The region encoding the amino-terminal 210 amino acids was then amplified with a FLAG tag at the carboxyl terminus from pcDNA3.1-hABCB6-C50A/C120A-V5 or pcDNA3.1-hABCB6-C8S/C50A/C120A-V5 by using primers (sense, 5Ј-GCCATGGTGACTGTGGGCAACTACTG- CGAGGCCG-3Ј; antisense, 5Ј-CCTACTTATCGTCGTCATC- CTTGTAATCACGAAGTCCAGGGGCCCAGAG-3Ј) under the following conditions: heating at 94 °C for 4 min, 30 cycles of denaturation at 94 °C for 30 s, annealing at 56 °C for 30 s, and reaction at 68 °C for 1 min. Login to comment
63 The inserts were subcloned into pcDNA3 using HindIII/XbaI sites to obtain pcDNA3.1- hABCB6-C50A/C120AN1-210-FLAG or pcDNA3.1-hABCB6-C8S/C50A/C120AN1-210-FLAG constructs. Login to comment
217 To rule out the contribution of the two additional cysteines (Cys-120 and Cys-150) present in this segment, we then developed a chimeric expression plasmid encoding the amino-terminal 210 amino acids of ABCB6 fused to a carboxyl-terminal FLAG epitope where both Cys-120 and Cys-150 were substituted with Ala (ABCB6-C50A/C120AN1-210-FLAG). Login to comment
220 We next compared the electrophoretic migration of ABCB6-C50A/C120AN1-210-FLAG and the cysteine mutant ABCB6-C8S/C50A/C120AN1-210-FLAG in the presence versus absence of the reducing agent DTT (Fig. 6B). Login to comment
223 ABCB6-C50A/ C120AN1-210-FLAG exhibited three faster migrating bands in the absence of DTT (Fig. 6B). Login to comment
257 B, ABCB6-C50A/C120AN1-210-FLAG and ABCB6-C8S/C50A/C120AN1-210-FLAG were expressed in NIH3T3 cells, treated with N-ethylmaleimide to prevent spontaneous disulfide bonds, separated by SDS-PAGE with or without DTT, and analyzed by immunoblotting. Login to comment