Molecular Mechanisms for Sweet-suppressing Effect of Gymnemic Acids
Molecular Mechanisms for Sweet-suppressing Effect of Gymnemic Acids
复制标题
DOI：
10.1074/jbc.m114.560409
复制
发表时间：
2014-09-12
影响因子：
4.8
通讯作者：
Ninomiya, Yuzo
中科院分区：
文献类型：
--
作者：
Sanematsu, Keisuke;Kusakabe, Yuko;Ninomiya, Yuzo
健美酸是一种三萜糖苷，在人类中可以选择性地抑制对各种甜味物质的味觉反应，但在老鼠身上则不是。用伽马-环糊精(伽马-环糊精)冲洗舌头会减弱体操酸的这种甜味抑制作用。然而，人们对体操酸抑制甜味的分子机制以及体操酸与甜味受体和/或γ-CD之间的相互作用知之甚少。为了研究运动酸是否与人(H)甜味受体hT1R2+hT1R3直接相互作用，我们在瞬时转染的HEK293细胞中使用了甜味受体T1R2+T1R3的检测方法。与之前在人类和小鼠身上的研究类似，健美酸(100mU g/ml)抑制了异源表达hT1R2+hT1R3的HEK293细胞中[Ca+](I)对甜味化合物的反应，但在表达小鼠(M)甜味受体mT1R2+mT1R3的细胞中不抑制。用γ-CD冲洗HEK293细胞后，健美酸的作用迅速消失。利用人和小鼠甜味受体亚基和嵌合体的混合物种配对，我们确定hT1R3的跨膜区主要是体酸抑制甜味所必需的。对hT1R3跨膜区的定向突变表明，与裸子酸相互作用的部位具有共同的氨基酸残基，这决定了其对另一种甜味拮抗剂乳酸的敏感性。葡萄糖醛酸是健美酸的常见结构，也会降低对甜味化合物的敏感度。在我们的模型中，预计体操酸会对接到hT1R3跨膜区内的结合口袋。
Gymnemic acids are triterpene glycosides that selectively suppress taste responses to various sweet substances in humans but not in mice. This sweet-suppressing effect of gymnemic acids is diminished by rinsing the tongue with gamma-cyclodextrin (gamma-CD). However, little is known about the molecular mechanisms underlying the sweet-suppressing effect of gymnemic acids and the interaction between gymnemic acids versus sweet taste receptor and/or gamma-CD. To investigate whether gymnemic acids directly interact with human (h) sweet receptor hT1R2 + hT1R3, we used the sweet receptor T1R2 + T1R3 assay in transiently transfected HEK293 cells. Similar to previous studies in humans and mice, gymnemic acids (100 mu g/ml) inhibited the [Ca2+](i) responses to sweet compounds in HEK293 cells heterologously expressing hT1R2 + hT1R3 but not in those expressing the mouse (m) sweet receptor mT1R2 + mT1R3. The effect of gymnemic acids rapidly disappeared after rinsing the HEK293 cells with gamma-CD. Using mixed species pairings of human and mouse sweet receptor subunits and chimeras, we determined that the transmembrane domain of hT1R3 was mainly required for the sweet-suppressing effect of gymnemic acids. Directed mutagenesis in the transmembrane domain of hT1R3 revealed that the interaction site for gymnemic acids shared the amino acid residues that determined the sensitivity to another sweet antagonist, lactisole. Glucuronic acid, which is the common structure of gymnemic acids, also reduced sensitivity to sweet compounds. In our models, gymnemic acids were predicted to dock to a binding pocket within the transmembrane domain of hT1R3.