MODEL SYNTHETIC CHANNEL ASSEMBLIES

合成通道组件模型

• 批准号: 7227438
• 负责人: John M Tomich
• 金额: $ 24.92万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7227438
• 关键词: Acids; Affect; Amino Acid Substitution; Anions; Antibiotics; Apical; Binding; Calibration; Cell membrane; Charge; Chemicals; Chloride Ion; Chlorides; Cholesterol; Class; Computer Simulation; Condition; Conflict (Psychology); Cornea; Cysteine; Cystic Fibrosis; Data; Data Analyses; Dental Cavity Lining; Detergents; Development; Dimensions; Environment; Epithelial; Epithelial Cells; Epithelium; Equilibrium; Future; Glycine Receptors; Goals; Head; Hydrogen Bonding; Hydroxyl Radical; Investigation; Ion Transport; Ionophores; Ions; Knowledge; Laboratories; Lead; Length; Light; Lipid Bilayers; Lipids; Liposomes; Measures; Membrane; Membrane Lipids; Methods; Micelles; Modeling; Movement; Nature; Numbers; Object Attachment; Outcome Study; Peptide Synthesis; Peptides; Pharmaceutical Preparations; Phenotype; Phospholipids; Physiological; Positioning Attribute; Production; Property; Range; Rate; Reaction; Reagent; Research Personnel; Scanning; Silver Staining; Site; Solutions; Spinal Cord; Standards of Weights and Measures; Structure; Synthesis Chemistry; Testing; Therapeutic; Therapeutic Intervention; Threonine; Ultracentrifugation; Xenopus oocyte; analytical method; base; basolateral membrane; bone strength; chemical synthesis; crosslink; design; membrane assembly; membrane model; monolayer; monomer; peptide structure; programs; protein aminoacid sequence; research study; response; sedimentation equilibrium; size; small molecule; stoichiometry; synthetic peptide; theories

DESCRIPTION (provided by applicant): Epithelia acts as barriers to the movement of small molecules, including ions and drugs, between body compartments. Ions cross the epithelial apical and basolateral membranes via tightly regulated ion-specific transporters and channels. However, exogenously supplied drugs, particularly hydrophilic or charged compounds, are not transported to any appreciable extent across the epithelium. Our labooratory has developed 2 different classes of peptides that promote selective ion transport or modulate the epithelial barrier that precludes drug access. The peptide sequences are derived from the pore-forming M2 transmembrane (TM) segment of the spinal cord glycine receptor (M2GlyR). Peptides insert and assemble to form transmembrane structures that display distinct, sequence dependent, physiological responses. They either selectively increase chloride transport or form non-selective channels that transiently facilitate the paracellular movement of larger hydrophilic compounds to the basolateral compartment. The former sequences have been proposed as a possible therapeutic intervention for channelopathies such as cystic fibrosis (CF). The latter are currently under investigation for facilitated transport of hydrophilic antibiotics across the cornea. While much is understood about the channel properties of these sequences in their native context little is understood about the assembled membrane structure(s) of the synthetic peptides. Multi-dimensional solution NMR has been used to investigate the structure of peptide monomers in SDS-micelles and modeling of helical bundles has been attempted. We propose to determine the number of transmembrane segments that assemble to form the active structures in model membranes and cells using both chemical and biophysical methods. Recent studies have indicated that different membrane lipid compositions can alter the secondary structure of these peptides. Studies will be conducted using model membranes with compositions that preseve the helical structure of the peptides. We hypothesize that different sized membrane assemblies could be stabilized in different lipid environments. Knowledge of the the number of segments that assemble under physiological conditions will greatly assist future modeling efforts. Additional experiments are planned that will examine the positioning and hydrogen bonding strength of a ring of threonines at position 17 in the sequences that form anion selective channel pores. Various amino acid substitutions in this position will assess the geometry and hydrogen bonding strengths required to optimize a synthetic channel with regard to selectivity and high throughput. These results will be incorporated into future channel designs with the ultimate goal of defining therapeutic structures.

描述（由申请人提供）：上皮细胞充当小分子（包括离子和药物）在身体区室之间移动的屏障。离子通过严格调节的离子特异性转运蛋白和通道穿过上皮顶膜和基底外侧膜。然而，外源提供的药物，特别是亲水性或带电化合物，不会以任何明显的程度跨上皮转运。我们的实验室开发了 2 种不同类别的肽，可促进选择性离子转运或调节阻止药物进入的上皮屏障。肽序列源自脊髓甘氨酸受体 (M2GlyR) 的成孔 M2 跨膜 (TM) 片段。肽插入并组装形成跨膜结构，显示出独特的、序列依赖性的生理反应。它们选择性地增加氯离子转运或形成非选择性通道，暂时促进较大亲水性化合物向基底外侧室的细胞旁运动。前一种序列已被提议作为囊性纤维化 (CF) 等通道病的可能治疗干预措施。目前正在研究后者促进亲水性抗生素穿过角膜的运输。虽然人们对这些序列在其天然背景下的通道特性了解很多，但对合成肽的组装膜结构了解甚少。多维溶液核磁共振已被用来研究 SDS 胶束中肽单体的结构，并尝试了螺旋束的建模。我们建议使用化学和生物物理方法确定模型膜和细胞中组装形成活性结构的跨膜片段的数量。最近的研究表明，不同的膜脂成分可以改变这些肽的二级结构。研究将使用具有保留肽螺旋结构的成分的模型膜进行。我们假设不同尺寸的膜组件可以在不同的脂质环境中稳定。了解在生理条件下组装的片段数量将极大地帮助未来的建模工作。计划进行额外的实验，检查形成阴离子选择性通道孔的序列中位置 17 的苏氨酸环的定位和氢键强度。该位置的各种氨基酸取代将评估在选择性和高通量方面优化合成通道所需的几何形状和氢键强度。这些结果将被纳入未来的通道设计中，最终目标是定义治疗结构。