Novel Organophosphorous Hydrolases for Decontamination

用于净化的新型有机磷水解酶

• 批准号: 6834554
• 负责人: ANDREW D MESECAR
• 金额: $ 9.62万
• 依托单位: LYBRADYN, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-20 至 2005-02-19
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6834554
• 关键词: bacterial proteins; bioreactors; biotechnology; bioterrorism /chemical warfare; biotransformation; directed evolution; environmental toxicology; enzyme activity; enzyme structure; esterase; gram positive bacteria; hazardous substances; mass tissue /cell culture; nerve gas; neurotoxins; organophosphorus insecticide; technology /technique development; transfection /expression vector

DESCRIPTION (provided by applicant): Organophosphorous (OP) neurotoxins such as paraoxon, sarin, soman, VX, and Russian-VX, are a broad class of chemicals commonly used as pesticides and chemical warfare (CW) agents. Currently, over 100,000 tons of chemical warfare agents are stockpiled worldwide, and thousands-of-tons of OP pesticides are released into the environment annually. Enzymes known as phosphotriesterases, commonly called organophosphorous hydrolases (OPH), are capable of hydrolyzing OP and CW agents into harmless products. Because of their potential for destruction of these neurotoxins, organophosphorous hydrolases have recently become the focus of intense research efforts aimed at developing these enzymes into products for enzyme-based decontamination. However, these enzymes lack many of the desired properties such as solution solubility, thermal and pH stability, and substrate specificity for all of the potential applications of these enzymes. We have recently identified, cloned, expressed, purified, crystallized, and determined the x-ray structure of a novel OPH from an extremophile that has significant activity against the chemical warfare agent soman. The enzyme can be overexpressed in large quantities, and has desirable physical properties such as high solubility and thermostability compared to all of the other OPH enzymes under development. Therefore, our version of OPH holds significant promise for technological advancement in both civilian and military applications that require novel OPH enzymes for enzyme-based decontamination. Since large-scale expression and production using our current expression vectors and host strains would not be economically viable, and since our version of OPH has low activity toward V-agents, we propose to develop an economically viable, industrial scale production system for D. radiodurans OPH enzymes, and to enhance the catalytic potential of D. radiodurans OPH towards V-agents and other organophosphorous compounds using directed evolution genetic technology.

描述（由申请人提供）：有机磷（OP）神经毒素，例如副毒素，沙林，索曼，VX和俄罗斯-VX，是一类广泛的化学物质，通常用作农药和化学战（CW）代理。目前，全世界已储存了100,000吨化学战的代理商，每年将数千千万的OP农药释放到环境中。称为磷酸三酯酶的酶，通常称为有机磷酸水解酶（OPH），能够将OP和CW剂水解为无害产物。由于它们可能会破坏这些神经毒素，因此有机磷液水解酶最近已成为旨在将这些酶开发为基于酶去污的产品的强烈研究工作的重点。但是，这些酶缺乏许多所需的特性，例如溶液溶解度，热和pH稳定性以及这些酶的所有潜在应用的底物特异性。我们最近已经确定，克隆，表达，纯化，结晶，并确定了来自对化学战剂Soman具有重要活性的新型OPH的X射线结构。与正在开发的所有其他OPH酶相比，该酶可以大量过表达大量的物理特性，例如高溶解度和热稳定性。因此，我们版本的OPH版本在需要新颖的OPH酶来进行基于酶的净化的新型OPH酶方面对技术进步具有巨大的希望。由于使用我们当前的表达向量和宿主菌株的大规模表达和生产在经济上是不可行的，并且我们的OPH版本对V-Agents的活性较低，因此我们建议为D. radiodurans OPH酶开发一种经济可行的，工业尺度的生产系统，以增强对远射型植物型植物的催化潜力，以增强对植物植物的催化剂的催化潜力。