Ultrasensitive Photoamplifed Fluorescence Detection of Ligand Binding on a Chip

描述(申请人提供)：分子识别事件的检测一直是生物分析科学的主要关注领域，因为生物分子对象与细胞中其他分子实体之间的结合在分子水平上支配着所有细胞功能。检测生物分子之间相互作用的能力对于开发新的治疗方法和治疗剂至关重要。这项提议的目标是开发一种全新的方法来对生物分子识别事件进行超灵敏放大检测，并将其应用于低成本的生物分析微阵列芯片。拟议的方法--基于电子转移敏感剂的自我放大揭幕--是标准化和普适性的，即不限于选定的生物分子和相互作用类别。一旦使用已知的配体-受体对进行验证，这项技术将被应用于分子生物学中的一个突出问题：识别组蛋白尾部的翻译后修饰。染色质领域中一个至关重要的问题是识别能够阅读组蛋白密码的新结构域。鉴于组蛋白尾部有大量的翻译后修饰，主要来自其赖氨酸和精氨酸残基的甲基化或乙酰化，寻找新的阅读器结构域只能通过高通量的组合筛选来实现。将合成包含天然组蛋白尾部签名残基的七肽文库，将其打印在放大的检测微阵列芯片上，并筛选已知的“读取”组蛋白编码的结构域--色域、溴域和PhD手指的结合。公共卫生相关性生物分子之间相互作用的检测一直是生物医学科学的主要关注领域，因为这种相互作用支配着生物有机体的所有过程。检测生物分子之间结合的能力对于开发新的治疗方法和治疗剂至关重要。这项提议的目标是开发一种低成本超灵敏检测方法的根本新方法，这将有助于发现新药。

DESCRIPTION (provided by applicant): Detection of molecular recognition events has always been an area of primary focus in bioanalytical sciences, as binding between biomolecular objects and other molecular entities in cells governs all cellular functions at the molecular level. The ability to detect interactions between biological molecules is vitally important for the development of new therapies and therapeutic agents. The goal of this proposal is to develop a fundamentally new approach to ultra-sensitive amplified detection of biomolecular recognition events and implement it in low cost bioanalytical microarray chips. The proposed methodology - which is based on self-amplified unmasking of electron-transfer sensitizers - is standardized and universal, i.e. not limited to selected classes of biological molecules and interactions. Once validated using known ligand-receptor pairs, this technology will be applied to a prominent problem in molecular biology: recognition of posttranslational modifications of histone tails. A critically important problem in the chromatin field is to identify new domains capable of reading the histone code. Given a large number of posttranslational modifications in histone tails, derived primarily from methylation or acetylation of their lysine and arginine residues, the search for new reader domains can only be achieved with high throughput combinatorial screening. Heptapeptide libraries containing signature residues of natural histone tails will be synthesized, printed on the amplified detection microarray chips, and screened for binding of domains known to "read" the histone code - chromodomain, bromodomain and the PHD finger. PUBLIC HEALTH RELEVANCE Detection of interactions between biological molecules has always been an area of primary focus in biomedical sciences, as such interactions govern all processes in living organisms. The ability to detect binding between biological molecules is vitally important for the development of new therapies and therapeutic agents. The goal of this proposal is to develop a fundamentally new approach to low cost ultra-sensitive detection methods, which will aid in the discovery of new medicines.