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Molucular analysis of tissue distribution and structure-function relationship of transporters responsible for the regulation of drug transport

负责药物转运调节的转运蛋白的组织分布和结构功能关系的分子分析

基本信息

批准号：
08457620
负责人：
INUI Ken-ichi
金额：
$ 5.57万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

Tissue distribution and structure-function relationship of membrane transporters responsible for absorption and excretion of drugs have been studied, and then following results were obtained.1.Structure-function relationship of H^+-coupled peptide transportersWhen stable transfectans expressing rat peptide transporters PEPT1 and PEPT2 were treated with DEPC, a histidine-modifying agent, glycylsarcosine uptake by both transfectants were decreased. Interactions of dipeptides and beta-lactam antibiotics were analyzed, and then it became clear that alpha-amino moiety of beta-lactam antibiotics should interact with histidine residues, and may participate in substrate recognition.2.Tissue distribution and functional characteristics of renal organic ion transporters.(1)Organic anion transporter, OAT-K1 : Expression of mRNA along nephron segments was analyzed by RT-PCR.OAT-K1 mRNAs were mainly expressed in proximal straight tubules of superficial and juxtamedullary nephrons. Western blot analysis revealed that OAT-K1 is expressed only in brush-border membranes of renal tubules. Methotrexate transport by the transfectants stably expressing OAT-K1 was significantly inhibited in the presence of nonsteroidal anti-inflammatory drugs (NSAID).(2)Organic cation transporter, OCT2 : Because tetraethylammonium transport by OCT2 was H^+-gradient independent and was affected by membrane potential, OCT2 is deduced to be basolateral-type organic cation transporter. By constructing stable transfectants expressing OCT1 or OCT2, transport characteristics were further analyzed, and then it became clear that both OCT1 and OCT2 are basolateral-type organic cation transporters with broad substrate specificity and have similar substate recognition in each other.

研究了负责药物吸收和排泄的膜转运蛋白的组织分布和结构-功能关系。1.H~+偶联多肽转运蛋白的结构-功能关系当表达大鼠肽转运蛋白PEPT1和PEPT2的稳定转运蛋白用组氨酸修饰剂DEPC处理时，这两种转染体对甘氨酰肌氨酸的摄取均减少。2.肾有机离子转运体的分布和功能特征：(1)有机阴离子转运体OAT-K1：RT-PCRNAs分析了OAT-K1在肾单位节段的表达。OAT-K1mRNAs主要表达于肾浅层和延髓旁肾单位的近端直管。Western印迹分析表明，OAT-K1仅在肾小管刷状缘膜中表达。在非甾体抗炎药(NSAID)存在下，稳定表达OAT-K1的转染体对甲氨蝶呤的转运受到明显抑制。(2)有机阳离子转运体OCT2：由于OCT2对四乙基铵的转运不依赖于H~+梯度，并且受膜电位的影响，推测OCT2是碱侧型有机阳离子转运体。通过构建稳定表达OCT1和OCT2的转染体，进一步分析了OCT1和OCT2的转运特性，发现OCT1和OCT2都是底物专一性较广的碱侧型有机阳离子转运体，具有相似的亚态识别能力。