STRUCTURE/FUNCTION OF OCULAR TRANSPORTERS

眼转运蛋白的结构/功能

• 批准号: 6329527
• 负责人: JORGE FISCHBARG
• 金额: $ 32.3万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-01 至 2001-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6329527
• 关键词: Xenopus oocyte; aminoacid; ascorbate; cell water; chemical kinetics; diffusion; edema; gene expression; glucose transport; interocular transfer; membrane permeability; membrane proteins; molecular dynamics; mutant; potassium channel; protein folding; protein structure function; site directed mutagenesis; sugar acids; water; water channel

DESCRIPTION: The PI has reported that both facilitative and Na+ dependent glucose transporters may transport their substrates through an aqueous pore. In addition, he has found that facilitative glucose transporters constitute the route of dehydroascorbic acid entry into cells, which may lead to a better understanding of the mechanisms that protect ocular tissues from oxidative damage. Glucose transporters appear to be gated channel proteins. Since solute transporters are essential to ocular function, he proposes to continue characterizing the structure-function relationship of facilitative glucose transporters (GLUTs) and other membrane proteins as archetypes of a number of solute transporters. He thus proposes: 1) To study GLUT structure by: a) cysteine scanning mutagenesis; function of mutants will be assayed by expression in oocytes; b) by 3-D modeling of proposed folds for GLUT1 such as the alpha-helical, beta-barrel, and mixed alpha-beta. We also propose to explore possible homologies between GLUT1 and a glucose transporter of known structure, maltoporin, as well as with segments of well-studied channel proteins such as aquaporins and potassium channels. 2) To investigate whether the passage of sugars and dehydroascorbic acid through GLUT1 can be ascribed to aromatic interactions with the transporter, in which case mutations that replace aromatic residues in GLUTs might affect the transporter selectivity and kinetics. 3) To characterize indirectly the GLUT channel cross section and binding properties by determining if molecules other than sugars traverse them, we have uncovered evidence that some amino acids (tyrosine, phenylalanine, leucine) traverse GLUT1. The PI proposes to characterize the kinetic parameters for each uptake mechanism, their sensitivity to inhibitors, and the degree to which these substrates can compete with each other, and with glucose and dehydroascorbate uptake. The hydrophobic envelopes of GLUT substrates will be determined by molecular dynamics simulations; this information will be used to deduce the shape and environmental characteristics of GLUT binding sites for these substrates. 4) To continue exploring the geometry and relative hydrophilicity of water permeable pathways in membrane proteins by comparing their water permeation characteristics with those obtained from molecular dynamics simulations and theoretical kinetic analysis of model channels.

描述：PI报告说促进和Na+依赖性 葡萄糖转运蛋白可以通过水孔运输其底物。 此外，他发现促进葡萄糖转运蛋白构成 脱氢抗坏血酸进入细胞的途径，这可能导致 更好地了解保护眼组织免受的机制 氧化损伤。 葡萄糖转运蛋白似乎是门控通道蛋白。 由于溶质转运蛋白对眼功能至关重要，他建议 继续表征促进的结构功能关系 葡萄糖转运蛋白（GLUT）和其他膜蛋白作为A的原型 溶质转运蛋白的数量。 他因此提出： 1）通过：a）半胱氨酸扫描诱变研究；功能 突变体的表达将在卵母细胞中测定； b）通过3-D建模 建议用于Glut1的折叠，例如α-螺旋，β-桶和混合 α-beta。 我们还建议探索Glut1之间可能的同源 以及已知结构的葡萄糖转运蛋白，以及 研究良好的通道蛋白（例如水通道蛋白和钾）的片段 频道。 2）研究糖和脱氢抗坏血酸是否通过 通过Glut1可以归因于转运蛋白的芳香相互作用， 在这种情况下，替代浮躁中芳香残基的突变可能会影响 转运蛋白的选择性和动力学。 3）间接表征Glut通道横截面并结合 通过确定除糖以外的分子是否穿越它们的特性，我们 有发现的证据表明一些氨基酸（酪氨酸，苯丙氨酸， 亮氨酸）遍历glut1。 PI提议表征动力学 每种摄取机制的参数，它们对抑制剂的敏感性，以及 这些基材可以彼此竞争的程度以及 葡萄糖和脱氢剂摄取。 疏水性包膜 底物将通过分子动力学模拟确定；这 信息将用于推断形状和环境 这些底物的过剩结合位点的特征。 4）继续探索水的几何形状和相对亲水性 通过比较膜蛋白的渗透途径 从分子动力学模拟获得的特征和 模型通道的理论动力学分析。