The expression of human catalytic proteins in micro algae on a commercial scale

利用人体蛋白质清除有机磷农药是一种很有前途的医学干预手段 用于预防和接触化学战神经毒剂后的治疗。最好的-研究 迄今为止，在临床前研究中取得了相当大的成功的生物清除剂（BSC）是人类的 胆碱酯酶（ChE）。然而，在结合和螯合OP方面非常有效的ChE也是 被毒素灭活，因此需要给予大量的蛋白质， 保护，提出了这种做法的实用性问题。改进型生物催化剂的发展 (BCT)可以催化降解OP的对氧磷酶（对氧磷酶1）可以解决这个问题。 所提出的努力提供了一种新的手段，以生物制造重组BSC和BCT的基础上， 人类蛋白质、丁酰胆碱酯酶和对氧磷酶1。与其他项目合作 参与该中心，这些蛋白质螯合或水解OP的能力将得到提高， 使它们的基因进行随机的体外进化或合理的诱变。为此，我们将 进行突变酶文库的高通量筛选，特别是提高稳定性， 对氧磷酶I的类药物性质。此外，翻译后糖基化系统或蛋白质 将开发聚乙二醇化程序，以增加循环寿命并消除潜在的 在非人重组生物体中产生的蛋白质的抗原性。OSU团队（项目6）将 利用转基因微藻，莱茵衣藻，生产中试规模的数量， 从培养基中直接回收的产品。此外，他们还将优化这一生产系统， 扩大到商业生产能力。该项目的主要意义在于， 用于生产增强的、第二代的、改进的基于胆碱酯酶的BSC和BCT的中试系统 适合人体临床试验的产品。

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. The development of improved biocatalysts (BCT) (paraoxonase 1) that can catalytically degrade OPs may address this concern. The proposed effort offers a novel means to biomanufacture recombinant BSCs and BCTs based on the human proteins, butyrylcholinesterase and paraoxonase 1. In collaboration with the other projects participating in the Center, the capacity of these proteins to sequester or hydrolyze OPs will be improved by subjecting their genes to either random in vitro evolution or rational mutagenesis. To that end, we will undertake high-throughput screening of mutant enzyme libraries, in particular to improve the stability and drug-like properties of paraoxonase I. In addition, post-translational glycosylation systems or protein PEGylation procedures will be developed to increase the circulating lifetimes and to eliminate potential antigenicity of proteins produced in non-human, recombinant organisms. The OSU team (Project 6) will utilize the transgenic microalgae, Chlamydomonas reinhardtii, to produce pilot-plant scale quantities of these products for direct recovery from the culture media. In addition, they will optimize this production system for scale-up to commercial production capacity. The primary significance of this project is that it will develop pilot-plant systems for the production of enhanced, second-generation, improved ChE-based BSC and BCT products suitable for human clinical trials.