Enhancement of organophosphinate hydrolase activity therough mechanistic evaluati

使用酶作为催化人类治疗剂对抗有机磷（OP）神经的关键障碍 代理商是他们的活性和特异性差。利用计算、机械、光谱和蛋白质 突变的方法，我们提出了生产和优化突变体的人类酶 （对氧磷酶I、丁酰胆碱酯酶和乙酰胆碱酯酶）催化化学物质的水解 这些神经毒剂的活性和特异性比迄今为止发现的更强。自从变异后 BuChE、AChE和HuPONI基于人类蛋白质，预期这些蛋白质将具有 很少或没有免疫学和行为副作用，使得这些合理的人类治疗药物 候选人我们将阐明潜在的化学作用机制所必需的催化 水解化学战神经毒剂，设计具有增强活性的人类蛋白质突变体 神经毒剂，并适当地设计人类来源的重组蛋白，然后可以 以足够的量表达用于随后的体内功效验证。总体预期是， 开发足够的科学数据，以便选择一种或两种蛋白质产品 作为新一代预防性生物制剂向高级开发过渡 可能被授予NDA地位，这些生物制剂将提供增强的保护， 神经毒剂中毒在军事或民用环境。计算的，机械的，光谱的， 光亲和标记、质谱、蛋白质组学和生物化学工具将用于野生型和 重组突变形式的胆碱酯酶和酯酶，用于开发这些新的 有机磷（OP）神经毒剂的治疗剂。 这些新形式的胆碱酯酶的发展将提供一种生物治疗方法 禁止在军事和民用环境中使用有机磷神经毒剂。这些疗法， 由于是人源的，将具有所需的化学特异性， 行为副作用

A critical barrier to using enzymes as catalytic human therapeutics against organophosphorous (OP) nerve agents is their poor activity and specificity. Using computational, mechanistic, spectroscopic, and protein mutagenic approaches, we propose the production and optimization of mutants of human enzymes (paraoxonase I, butyrylcholine esterase, and acetylcholinesterase) to catalyze the hydrolysis of chemical warfare nerve agents with greater activity and specificity than those identified to date. Since mutated BuChE, AChE and HuPONI are based on human proteins, the expectation is that these proteins would have few or no immunological and behavioral side effects, making these reasonable human therapeutic candidates. We will elucidate the underlying chemical mechanisms of action necessary for catalytic hydrolysis of chemical warfare nerve agents, design mutants of human proteins with enhanced activity toward nerve agents, and appropriately design recombinant proteins of human origin which can then be expressed in sufficient quantities for subsequent in vivo validation of efficacy. The overall expectation is to develop a sufficient body of scientific data to allow for a selection to be made of one or two protein products for the transition to advanced development as a new generation of prophylactic biological agents with the potential to be granted NDA status and that these biological agents will provide enhanced protection against nerve agent poisoning in a military or civilian setting. Computational, mechanistic, spectroscopic, photoaffinity labeling, mass spectrometric, proteomic, and biochemical tools will be used with wild-type and recombinant mutant forms of cholinesterase and esterase enzymes for the development of these novel therapeutics against organophosphorous (OP) nerve agents. The development of these novel forms of cholinesterase enzymes will provide a biological therapeutic against the use of organophosphorous nerve agents in military and civilian settings. These therapeutics, being of human origin, will have the desired chemical specificity and also few or no immunological and behavioral side effects