Alterations in the primary structure of the active transport carrier and changes in the function

主动转运载体一级结构的改变和功能的变化

• 批准号: 61580145
• 负责人: TSUCHIYA Tomofusa
• 金额: $ 1.28万
• 依托单位: Okayama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1986
• 资助国家: 日本
• 起止时间: 1986 至 1987
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-61580145/
• 关键词: Transport carrier; Primary structure; Amino acid substitution; Changes in function; カチオン共役; 一次構造の変化; 機能の変化; 変異の同定

Several transport systems for nutrients in cell membranes are Na^+-coupled cotransport systems. One of the best characterized Na^+-coupled cotransport systems is the melibiose transport system in Esherichia coli.We isolated many mutants which showed altered cation specificity in the melibiose carrier. There were two types of mutant. The melibiose carrier of one type of the mutants lost the ability to couple to H^+ and acquired the ability to couple to Li^<> when melibiose was the substrate. The coupling to Na^+ in those mutants was normal. The melibiose carrier of another type of the mutants lost the ability to couple to H^<> and could utilize only Na^+ as the coupling csation. We analysed the second type mutants in this project.Mutated gene, melB, of the mutants was cloned, and the nucleotide sequence was determined. The nucleotide replacements caused the following substitutions of amino acid residues in the melibiose carrier; Pro-142 with Ser, Leu-232 with Phe, or Ala-236 with Thr or Val. These amino acid residues are located in slightly hydrophobic regions of the melibiose carier. The results provide a strong support for the idea that such regions or their vicinities which contain those amino acid residues play an important role in H^+ recognition or H^+ transport by the melibiose carrier.

细胞膜中营养物质的几种运输系统是钠离子偶联的共运输系统。在大肠杆菌中，Na~+偶联共转运系统是最具特性的Na~+偶联共转运系统之一。我们分离到了许多突变株，它们在该载体上表现出阳离子特异性改变。有两种类型的突变体。其中一种突变株的蜜二糖载体失去了与H^+偶联的能力，而在以蜜二糖为底物时获得了与Li^&gt；的偶联能力。这些突变体与Na+的偶联是正常的。另一类突变体的杂二糖载体失去了与H^&lt；&gt；的偶联能力，只能利用Na~+作为偶联离子。对本项目中的第二类突变体进行了分析，克隆了突变体的突变基因Melb，并测定了其核苷酸序列。核苷酸替换导致美二糖载体中的氨基酸残基发生以下替换：Pro-142与Ser，Leu-232与Phe，或Ala-236与Thr或Val。这些氨基酸残基位于Melibiose carier的轻微疏水性区域。这些结果有力地支持了这一观点，即含有这些氨基酸残基的这些区域或邻近区域在蜜二糖载体的H^+识别或H^+运输中起着重要作用。

项目成果

石川哲也: Journal of Biological Chemistry. 262. 7443-7446 (1987)

石川哲也：生物化学杂志 262. 7443-7446 (1987)

Dorothy M.Wilson: Methods in Enzymology. 125. 377-387 (1986)

多萝西·M·威尔逊：酶学方法。

堺由妃: Biochemical and Biophysical Research Communication. 144. 382-386 (1987)

Yuki Sakai：生物化学和生物物理研究通讯。144。382-386（1987）

Sakai,Yuki: "A novel mechanism for utilization of extracellular AMP in Vibrio Parahaemolyticus" Biochemical and Biophysical Research Communications. 144. 382-386 (1987)

Sakai，Yuki：“副溶血弧菌利用细胞外 AMP 的新机制”生物化学和生物物理研究通讯。

濱啓子: Biochimica et Biophysica Acta. 905. 231-239 (1987)

Keiko Hama：生物化学与生物物理学学报 905. 231-239 (1987)