Mutually Exclusive Protein Folding

描述（由申请人提供）：该项目的目标是开发蛋白质折叠与展开耦合的机制，以便将变构控制引入蛋白质和酶。为此介绍了两种技术：互斥折叠（MEF）和交替框架折叠（AFF）。MEF利用储存在一个区域折叠结构中的自由能来展开同一分子内的另一个区域。这两个领域不可能同时以各自的状态存在；因此分子在两种功能形式之间相互转化。通过将受体结构域与酶或功能结构域相匹配，可以创造出新的分子，其中配体结合可以产生所需的生物功能。AFF是一种以简单、可预测和定义良好的方式耦合结合构象变化的新机制。它包括氨基酸序列的排列和部分序列的重复。由此产生的蛋白质在结构相似但具有不同拓扑结构的天然结构之间切换。动力是由配体结合和蛋白质折叠之间的普遍联系提供的。构象变化被用于生物功能或输出信号。AFF可以在单个蛋白内实现，也可以与MEF在双域融合中结合以建立变构调节。其目标是产生用于钙和糖的生物传感器，用于开发多功能（“通用”）传感器的蛋白质支架，一种有效的抗hiv毒素，以及一种将条件功能引入选择蛋白质的方法。这些分子将为设计具有新型分子识别和治疗特性的各种蛋白质建立一个框架。

DESCRIPTION (provided by applicant): The goal of this project is to develop mechanisms by which protein folding can be coupled to unfolding, in order to introduce allosteric control into proteins and enzymes. Two techniques are introduced for this purpose: mutually exclusive folding (MEF) and alternate frame folding (AFF). MEF uses the free energy stored in the folded structure of one domain to unfold another domain within the same molecule. The two domains cannot exist simultaneously in their native states; hence the molecule interconverts between two functional forms. By matching a receptor domain with an enzymatic or functional domain, one can create new molecules in which ligand binding causes a desired biological function. AFF is a novel mechanism for coupling binding to conformational change in a simple, predictable, and well-defined manner. It involves permutation of the amino acid sequence and partial sequence duplication. The resulting protein switches between native structures that are structurally similar, but possess different topologies. The driving force is provided by the universal linkage between ligand binding and protein folding. The conformational change is harnessed to a biological function or to an output signal. AFF can be implemented within a single protein or can be combined with MEF in a two- domain fusion to establish allosteric regulation. The aims will generate biosensors for calcium and sugars, protein scaffolds for development of versatile ("universal") sensors, a potent anti-HIV toxin, and a method for introducing conditional functionality to a protein of choice. These molecules will establish a framework for designing a wide variety of proteins with novel molecular recognition and therapeutic properties. PUBLIC HEALTH RELEVANCE: This project develops new mechanisms for introducing allosteric control into proteins. This technology is used to create proteins of medical significance (anti-HIV therapeutics) and biological interest (biosensors for metals, sugars, and other small molecules).