PRODUCTION OF ALPHA-GALACTOSIDASE A--TRANSFECTED PLANTS

成功开发负担得起的分子疗法将需要 同时生产和生物加工的全新方法 重组蛋白。例如，新陈代谢储存障碍是 最常见的一组遗传异常和许多这样的情况 很可能是可以通过补充外源生产的 酵素。然而，对于任何特定的分子缺陷，少量的 个人受到影响，因此预计治疗费用非常高 很高。Fabry病是一种分解代谢中的隐性X连锁缺陷 溶酶体水解酶α-半乳糖苷酶A(GA1-A)和IS的活性 人类第二常见的遗传性代谢储存障碍。这个 这项建议的总体目标是确定 经济地生产用于治疗的定制设计的重组GA1-A 治疗法布里病。人类来源的GA1-A具有同源二聚体 亚基分子量约为50的糖蛋白结构 KDA。在最初的实验中，我们将GalA基因克隆到病毒载体中 并确定了生产和提纯的技术可行性。 从转基因植物的叶片中分离出这种酶。产率、纯度和 酶的质量保证了进一步的生化和结构分析 评估其在人体内输血的适宜性。 建议的商业应用： 这项技术将建立工厂作为一项重要而经济的 大量重组蛋白的来源 推进现代分子疗法。成本优势基于 病毒的快速导入，产物的有利水平 粗提物中的浓缩，以及从 植物生物质，一种可再生、低投入和易于扩展的来源。

Successful development of affordable molecular therapies will require fundamentally new approaches to simultaneous production and bioprocessing of recombinant proteins. For example, metabolic storage disorders are the most common group of hereditary abnormalities and many of these conditions are likely to be treatable by supplementation with exogenously produced enzymes. Yet for any particular molecular defect, small numbers of individuals are affected and therefore projected treatment costs are very high. Fabry disease is a recessive X-linked deficiency in the catabolic activity of the lysosomal hydrolase alpha-galactosidase A (Ga1-A) and is the second most common hereditary metabolic storage disorder of man. The overall objective of this proposal is to determine the feasibility of economically producing custom-designed recombinant Ga1-A for therapeutic treatment of Fabry disease. Ga1-A from human sources has a homodimeric glycoprotein structure with a subunit molecular weight of approximately 50 kDa. In initial experiments, we cloned the Gal-A gene into a viral vector and established technical feasibility for the production and purification of this enzyme from transfected plant leaves. The yield, purity and quality of the enzyme warrant further biochemical and structural analyses to evaluate its suitability for transfusions into human subjects. PROPOSED COMMERCIAL APPLICATION: This technology will establish plants as an important and economical source of the large quantities of recombinant proteins necessary to advance modern molecular therapies. Cost advantages are based on the rapidity of the viral transfection, the favorable levels of product enrichment in crude extracts, and the high yield of product obtained from plant biomass, a renewable and low-input and easily scalable source.