Rapid and large scale plant-based production of catalytic nerve agent bioscavenge

利用人类蛋白质清除有机磷(OP)正成为一种很有前途的医学干预手段 用于预防和暴露后治疗化学战神经毒剂。研究最多的人 迄今为止，生物清洁剂(BSC)在临床前研究中取得了相当大的成功，它们都是人类 胆碱酯酶(Ches)。然而，在绑定和隔离OP方面非常高效的CHE也是 被毒素灭活，因此需要大量的蛋白质才能完全 保护，提出了这一办法的实用性问题。然而，一种新的 能够催化降解OP的BSC的产生可能会解决这一问题。 该工作提供了一种生物制造重组催化BSCs的新方法 人蛋白丁酰胆碱酯酶和对氧磷酶1.通过参与其他项目的努力 中心，这些蛋白质的能力将被提高，通过使他们的基因 要么是随机的体外进化，要么是理性突变。伴随着蛋白质工程研究， 在这个项目5中，亚利桑那州立大学的团队将利用烟草植物首先生产研究规模的BSC 酶，也提供了可持续的规模化生产平台。目前，从植物中分离纯化BChE的方法较多 过时的血库人类血浆使研究如何使用生物清道夫疗法成为可能。但这件事 不能实际实施权宜之计，以允许持续供应这种酶，而且 不适用于该中心将开发的新型重组催化BSCs。如果是这样的话 决定这种拾荒者应成为国防部医疗武器库的一部分， 国土安全部和卫生与公众服务部，至关重要的是，可靠、安全、非限制供应和 确定和开发这种酶的廉价来源。建议的主要意义 其工作是将第一代基于CHE的BSCs的基础研究转化为新的生物制造 引领第二代催化骨髓干细胞临床产品开发的技术。

Bioscavenging of organophosphate (OP) by human proteins is emerging as a promising medical intervention for prophylaxis and post-exposure treatment against chemical warfare nerve agents. The best-studied bioscavengers (BSCs) to date, meeting considerable success in pre-clinical research, are human cholinesterases (ChEs). However, ChEs, which are highly efficient in binding and sequestering OPs, are also inactivated by the toxins and therefore administration of large amounts of protein is necessary for full protection, raising the question of the practicality of this approach. However the development of a new generation of BSCs that can catalytically degrade OPs may address this concern. The proposed effort offers a novel means to biomanufacture recombinant catalytic BSCs based on the human proteins butyrylcholinesterase and paraoxonase 1. Through efforts of other projects participating in the Center, the capacity of these proteins to hydrolyze OPs will be improved by subjecting their genes to either random in vitro evolution or rational mutagenesis. Concomitantly to the protein engineering research, the ASU team in this Project 5 will utilize tobacco plants to first produce research-scale quantities of BSC enzyme, but also provide a sustainable large-scale production platform. At present, purification of BChE from outdated blood-banked human plasma enables research on how bioscavenger therapy can be used. But this stop-gap measure cannot be practically implemented to allow for a sustained supply of that enzyme, and is not applicable for the new recombinant catalytic BSCs that will be developed by the Center. If it will be decided that such scavengers should be a component of the medical arsenal of the Departments of Defense, Homeland Security and Health and Human Services, it is vital that a reliable, safe, non supply-limited and inexpensive source of such enzymes be identified and developed. The primary significance of the proposed work is that it translates basic studies on the first generation ChE-based BSCs into novel biomanufacturing technology leading to clinical product development of the second generation catalytic BSCs.