Model Synthetic Channel Assemblies

模拟合成通道组件

• 批准号: 8065348
• 负责人: John M Tomich
• 金额: $ 29.89万
• 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 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8065348
• 关键词: Acids; Amino Acids; Anions; Area; Binding; Bio-Base; Biological; Cations; Cells; Characteristics; Charge; Chemicals; Computer Simulation; Cysteine; Cystic Fibrosis; Diffusion; Dipeptides; Disease; Electrolytes; Electrophysiology (science); Electrostatics; Environment; Epithelial; Generations; Glycine Receptors; Goals; Half-Life; Hydrogen Bonding; Ion Channel; Ion Transport; Ions; Laboratories; Lead; Length; Life; Liquid substance; Membrane; Methods; Modeling; Modification; Monitor; Monovalent Cations; Morbidity - disease rate; Movement; Outcome; Parents; Peptide Synthesis; Peptides; Pharmaceutical Preparations; Positioning Attribute; Property; Proteins; Regulation; Relative (related person); Replacement Therapy; Series; Solubility; Solutions; Source; Spinal Cord; Structure; Study models; Technology; Therapeutic; Threonine; Tight Junctions; Water; Work; alanylproline; apical membrane; aqueous; basolateral membrane; crosslink; design; extracellular; flexibility; interfacial; loss of function; monolayer; monomer; mortality; protein aminoacid sequence; public health relevance; response; simulation; solute; structural biology; synthetic peptide

DESCRIPTION (provided by applicant): The goal of this project is to design, develop and implement the use of ion-selective, synthetic channel-forming peptides. A primary application for such technology will be the treatment of loss-of-function ion channel diseases such as cystic fibrosis. Epithelial monolayer's act as barriers to the movement of small solute molecules, including both inorganic ions and drugs, between body compartments. Ions cross epithelial apical and basolateral membranes via combination of tightly regulated ion-specific transporters and channels; limited diffusion across tight junctions can occur. Loss-of-function of any channel leads to electrolyte and fluid imbalances that result in morbidity and mortality. For many years, this laboratory has been developing synthetic peptides that form pores with varying degrees of anion conduction and selectivity. These synthetic peptide sequences are distantly related to the pore-forming transmembrane segment (M2) of the spinal cord glycine receptor (GlyR) a1-subunit. We hypothesized that the ideal channel-forming sequence should:1) have high aqueous solubility as a monomer; 2)have no detectable antigenicity;3) bind to and then partition rapidly into biological membranes at low solution concentrations; 4) undergo supramolecular assembly in the membrane to form pores with high ion throughput; and 5) show physiologically relevant anion selectivity. All of these properties have been achieved with the exception of the final goal, anion selectivity. We are now exploring distinct approaches to raise the permselectivity for Cl- (PCl) relative to either Na+ or K+. Cells treated with our de novopores tolerate them well with the net anion flux controlled by the natural regulation of counter-ion transport. In the Aims we propose to modulate pore-exposed and peptide-peptide interfacial residues that dictate the pore environment, channel geometry, and channel size to generate a series of pores that define a range of perm selectivity's for Cl- relative to monovalentcations, with retention of high anion permeation rates. Specific amino acid replacements will be introduced to modulate pore lining residues with regard to hydrogen bonding capabilities or electrostatic properties. Other replacements will alterpore length and rigidity. The effects of these modifications will be monitored by a combination of electrophysiological and structural (CD & NMR) studies in conjunction with computer modeling. PUBLIC HEALTH RELEVANCE: The bio-based peptide materials described in this proposal are derived from the same biological source and undergo self assemble in both membranes and living cells to form channel pores that display unique biological properties. Producing new peptides with high anion selectivity would have translational promise in the area of channel- replacement therapy for channelopathies such as cystic fibrosis.

描述（由申请人提供）：该项目的目标是设计、开发和实施离子选择性合成通道形成肽的使用。这种技术的主要应用是治疗囊性纤维化等功能丧失的离子通道疾病。上皮单层充当小溶质分子（包括无机离子和药物）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的组合穿过上皮顶膜和基底外侧膜；可能会发生跨紧密连接的有限扩散。任何通道的功能丧失都会导致电解质和液体失衡，从而导致发病和死亡。多年来，该实验室一直在开发合成肽，这些肽形成具有不同程度的阴离子传导和选择性的孔。这些合成肽序列与脊髓甘氨酸受体 (GlyR) a1 亚基的成孔跨膜片段 (M2) 关系较远。我们假设理想的通道形成序列应该：1）作为单体具有高水溶性； 2）没有可检测到的抗原性；3）在低溶液浓度下结合并快速分配到生物膜中； 4）在膜内进行超分子组装，形成高离子通量的孔； 5) 显示生理相关的阴离子选择性。除了最终目标（阴离子选择性）之外，所有这些特性均已实现。我们现在正在探索不同的方法来提高 Cl- (PCl) 相对于 Na+ 或 K+ 的选择性渗透性。用我们的 de novopore 处理的细胞能够很好地耐受它们，其净阴离子通量由反离子运输的自然调节控制。在目标中，我们建议调节孔暴露和肽-肽界面残基，这些残基决定孔环境、通道几何形状和通道尺寸，以生成一系列孔，这些孔定义了相对于单价阳离子的 Cl- 的一系列渗透选择性，并具有保留高阴离子渗透率。将引入特定的氨基酸替代物来调节孔衬里残基的氢键能力或静电特性。其他替代品会改变孔隙长度和刚性。这些修饰的效果将通过电生理学和结构（CD 和 NMR）研究与计算机建模相结合来监测。 公共健康相关性：本提案中描述的生物基肽材料源自相同的生物来源，并在膜和活细胞中进行自组装，形成显示独特生物特性的通道孔。生产具有高阴离子选择性的新肽将在囊性纤维化等通道病的通道替代疗法领域具有转化前景。