Improvement of glycosyltransferases for in vitro glycan remodeling of plant derived therapeutic proteins.

改进糖基转移酶，用于植物源性治疗蛋白的体外聚糖重塑。

• 批准号: 550399-2020
• 负责人: Wakarchuk, Warren
• 金额: $ 5.54万
• 依托单位: University of Alberta
• 依托单位国家: 加拿大
• 项目类别: Alliance Grants
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=701820
• 关键词: Improvement; glycosyltransferases; vitro; glycan; remodeling

Plants represent an excellent cost-effective alternative to mammalian cell culture for production of protein based drugs known as biologics. These drugs are proteins derived from normal human biology, where they protect us from various natural and man-made toxins, infectious disease, and immune system malfunctions like cancer. These are produced as recombinant proteins and are very expensive to manufacture in tissue culture. Normal human therapeutic proteins are decorated with molecules called glycans making them "glycoproteins", and these glycans are essential for stability and longevity in the body. The biochemical machinery for making glycoproteins in plants and animals is almost identical, resulting in plants being able to make some important human proteins, like antibodies, and proteins which protect from toxins like nerve gas. Plants however don't naturally make human style glycan decorations on plants, and plant glycans can cause an allergic reaction. Recent advances in plant engineering now allows researchers to produce proteins without the plant specific decorations which make the proteins a target of the human immune system. However, the production of the authentic human glycan decorations is still difficult to control in both mammalian- and plant-based expression systems. PlantForm, and the Wakarchuk lab are collaborating to use enzymes in the laboratory to finish the glycan decorations exactly as those found on the normal human protein with the goal of commercial-scale glycan modification of target proteins. To this end, the enzymes required to change the glycans are being engineered for improved production, activity and stability through our unique bacterial expression systems. We are developing immobilized enzymes to produce various terminal glycan decorations such that the proteins don't require extensive re-purification after the reactions. We are also engineering production of these enzymes in a bacterium which doesn't produce a toxic contaminant - endotoxin - which requires significant effort to remove from these high value proteins.

植物代表了一种极好的成本效益替代哺乳动物细胞培养生产蛋白质为基础的药物被称为生物制剂。这些药物是来自正常人类生物学的蛋白质，它们保护我们免受各种天然和人造毒素、传染病和免疫系统故障（如癌症）的侵害。它们以重组蛋白的形式产生，在组织培养中制造非常昂贵。正常的人类治疗性蛋白质被称为聚糖的分子修饰，使其成为“糖蛋白”，这些聚糖对人体的稳定和长寿至关重要。植物和动物制造糖蛋白的生化机制几乎完全相同，因此植物能够制造一些重要的人类蛋白质，如抗体，以及保护人体免受神经毒气等毒素侵害的蛋白质。然而，植物不会自然地在植物上制造人类风格的聚糖装饰，植物聚糖会引起过敏反应。植物工程的最新进展现在允许研究人员生产没有植物特异性修饰的蛋白质，这使得蛋白质成为人类免疫系统的目标。然而，在哺乳动物和植物的表达系统中，真正的人聚糖修饰物的产生仍然难以控制。PlantForm和Wakarchuk实验室正在合作，在实验室里使用酶来完成聚糖修饰，就像在正常人类蛋白质上发现的那样，目标是实现商业规模的聚糖修饰目标蛋白质。为此，通过我们独特的细菌表达系统，我们正在设计改变聚糖所需的酶，以提高产量、活性和稳定性。我们正在开发固定化酶，以产生各种末端聚糖修饰，使蛋白质在反应后不需要大量的再纯化。我们还在一种细菌中设计这些酶的生产，这种细菌不会产生有毒污染物——内毒素——从这些高价值的蛋白质中去除内毒素需要付出很大的努力。