title: ATPase activity of nucleotide binding domains of human MDR3 in the context of MDR1.
abstract: Although human MDR1 and MDR3 share 86% similarity in their amino acid sequences and are predicted to share conserved domains for drug recognition, their physiological transport substrates are quite different: MDR1 transports xenobiotics and confers multidrug resistance, while MDR3 exports phosphatidylcholine into bile. Although MDR1 shows high ATPase activity, attempts to demonstrate the ATPase activity of human MDR3 have not succeeded. Therefore, it is possible that the difference in the functions of these proteins is caused by their different ATPase activities. To test this hypothesis, a chimera protein containing the transmembrane domains (TMDs) of MDR1 and the nucleotide binding domains (NBDs) of MDR3 was constructed and analyzed. The chimera protein was expressed on the plasma membrane and conferred resistance against vinblastine and paclitaxel, indicating that MDR3 NBDs can support drug transport. Vanadate-induced ADP trapping of MDR3 NBDs in the chimera protein was stimulated by verapamil as was MDR1 NBDs. The purified chimera protein showed drug-stimulated ATPase activity like MDR1, while its Vmax was more than 10-times lower than MDR1. These results demonstrate that the low ATPase activity of human MDR3 cannot account for the difference in the functions of these proteins, and furthermore, that TMDs determine the features of NBDs. To our knowledge, this is the first study analyzing the features of human MDR3 NBDs.
authors.affiliation: Laboratory of Cellular Biochemistry, Division of Applied Life Sciences, Kyoto University Graduate School of Agriculture, Kyoto 606-8502, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Ishigami
authors.forename: Yuko
authors.initials: Y
authors.lastname: Tominaga
authors.forename: Kohjiro
authors.initials: K
authors.lastname: Nagao
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Kimura
authors.forename: Michinori
authors.initials: M
authors.lastname: Matsuo
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Kioka
authors.forename: Kazumitsu
authors.initials: K
authors.lastname: Ueda
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018432:Drug Resistance, Multiple; D006801:Humans; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism*; D017239:Paclitaxel / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014700:Verapamil / Q000494:pharmacology; D014747:Vinblastine / Q000494:pharmacology
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D009711:Nucleotides; D011994:Recombinant Proteins; D014747:Vinblastine; C095122:multidrug resistance protein 3; D014700:Verapamil; D000251:Adenosine Triphosphatases; D017239:Paclitaxel