Alteration of Substrate Specificity by Mutations at the His61 Position in Predicted Transmembrane Domain 1 of Human MDR1/P-Glycoprotein

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• 作者：Yoshitomo Taguchi ; Kouichi Kino ; Masaki Morishima ; Tohru Komano ; Susan E. Kane ; and Kazumitsu Ueda
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：July 22, 1997
• 年：1997
• 卷：36
• 期：29
• 页码：8883 - 8889
• 全文大小：251K
• 年卷期：v.36,no.29(July 22, 1997)
• ISSN：1520-4995

文摘

In CFTR, a member of the ABC superfamily and achloride channel, amino acid substitutionsin its transmembrane domains 1 and 6 (TM1, TM6) have been reported tomodulate the anion selectivityor ion conductance of the ion channel. In P-glycoprotein, no aminoacid substitution in TM1, but somein TM6, have been reported to modify the substrate specificity of thisprotein. In this work, wedemonstrated the involvement of His⁶¹, which is in themiddle of the predicted TM1, in the function ofP-glycoprotein. His⁶¹ was replaced by all other aminoacid residues, and each of the mutant cDNAs wasintroduced into drug-sensitive human carcinoma cells, KB3-1. Thedrug-resistance profile of cells stablyexpressing each mutated P-glycoprotein was investigated by comparingtheir relative resistance tovinblastine, colchicine, VP16, and adriamycin. The resistance tovinblastine was increased by replacingHis⁶¹ by amino acids with smaller side chains, while it waslowered by replacing by amino acids withbulkier side chains. The reverse effect was observed forresistance to colchicine and VP16. The resistanceto adriamycin was increased by replacing by amino acids with bulkierside chains except Lys or Arg,which have a basic side chain. We also showed that the replacementof His⁶¹ by Phe and Lys greatlyimpaired the efflux of calcein AM, while the replacement had no effecton the efflux of rhodamine 123.These results suggest that an amino acid residue at position 61 inTM1 is important in deciding the substratespecificity of P-glycoprotein.