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Plant-produced Asialo-erythropoietin and its Antiapoptotic Functions

植物性促红细胞生成素及其抗细胞凋亡功能

基本信息

批准号：
7905048
负责人：
Jiahua Xie
金额：
$ 10.5万
依托单位：
NORTH CAROLINA CENTRAL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-01 至 2013-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7905048
关键词：
Academia Adverse effects Advocate Amino Acids Animal Model Applications Grants Appointment Area Biological Process Biological Products Biomanufacturing Biomass Biomedical Engineering Bioreactors Biotechnology C-terminal Cardiac Myocytes Cardiotonic Agents Cell Culture Techniques Cell Death Cell Line Cells Codon Nucleotides Collaborations Communities Complex Data Development Discipline Educational process of instructing Erythrocytes Erythropoietin Faculty Foundations Fucose Funding Future Galactose Galactosyltransferases Gene Expression Genes Genetic Genetically Modified Plants Glycoproteins Goals Grant Harvest Hematopoietic Hormones Human Immunoblotting Industry Institutes Institution Kanamycin Resistance Knowledge Laboratories Left Link Mammalian Cell Mammals Measures Medical Methodology Methods Minority Mission Modification Molecular Molecular Biology Mus Neurons North Carolina Oxygen PC12 Cells Pattern Peptide Signal Sequences Pharmacologic Substance Plant Leaves Plants Play Polysaccharides Process Production Productivity Property Protein Chemistry Proteins Protocols documentation Publications Publishing Research Research Activity Research Institute Research Personnel Research Support Risk Role Science Seeds Sialic Acids Solid Structure Students System Techniques Technology Testing Tissues Tobacco Tobacco use Training Transgenic Organisms Transgenic Plants Universities Vision Work Xylose base career cost forest glycosylation hematopoietic tissue improved in vivo insight interest large scale production novel pathogen prevent professor protein purification public health relevance receptor recombinant human erythropoietin scale up segregation sugar symposium systems research vector

项目摘要

DESCRIPTION (provided by applicant): In this project, we are taking the distinct advantage of plant expression systems, which is the lack of the capacity to add sialic acids, to genetically modify plants for asialo-rhuEPO - a special glycoprotein without sialic acids. Although nearly all human glycoproteins require sialic acid residues for their in vivo functions, the one exception we could find is rhuEPO, which has both hematopoietic and tissue protective functions. When rhuEPO is used as an antiapoptotic agent, its hematopoietic activity can cause bad side effects by leading to harmful increases in red blood cell mass. This activity can be disrupted by removing sialic acids. Asialo-rhuEPO generated by total enzymatic desialylation of rhuEPO has been demonstrated to retain the cell- and tissue-protective properties of EPO without hematopoietic activity. This type of EPO derivative can be explored as the antiapoptotic agent. However, there is no expression system that can produce asialo-rhuEPO directly. It is also unlikely to use commercially available rhuEPO, which is produced only in mammalian cells, to make asialo-rhuEPO because of its limited supply. Based on available knowledge and techniques, it is likely to genetically modify plants to produce human glycoproteins without sialic acids. We have co- expressed human GalT gene and EPO gene fusing with ER-signal peptides in tobacco plants, and try to inexpensively produce asialo-rhuEPO with the potential of large scale production. We expect plant-produced asialo-rhuEPO will have similar cytoprotective function as the enzymatically produced asialo-rhuEPO. The results of this project will also provide insights into the plant glycosylation mechanism and the antiapoptotic mechanism of the plant- produced asialo-rhuEPO. In addition, the protein purification methodologies that will be established and the glycoprotein analysis methods will be very valuable for future plant glycoengineering projects. PUBLIC HEALTH RELEVANCE: We aim to genetically modify plants for production of asialo-rhuEPO - a special human protein. This type of biopharmaceutical protein can be explored as the neurotherapeutic and cardioprotective agent. Using a plant expression system, we expect to produce an inexpensive and functional asialo-rhuEPO to prevent cell death with the potential of large scale production.

描述（由申请人提供）：在本项目中，我们利用植物表达系统的独特优势，即缺乏添加唾液酸的能力，对植物进行无唾液酸-rhuEPO-一种不含唾液酸的特殊糖蛋白的遗传修饰。虽然几乎所有的人类糖蛋白都需要唾液酸残基来维持其体内功能，但我们可以找到的一个例外是rhuEPO，它具有造血和组织保护功能。当rhuEPO用作抗凋亡剂时，其造血活性可通过导致红细胞质量的有害增加而引起不良副作用。这种活性可以通过去除唾液酸来破坏。通过rhuEPO的总酶促去唾液酸化产生的脱唾液酸rhuEPO已被证明保留EPO的细胞和组织保护特性而没有造血活性。这种类型的EPO衍生物可以作为抗凋亡剂进行开发。然而，目前还没有一个能直接生产去唾液酸rhuEPO的表达系统。由于其供应有限，也不太可能使用仅在哺乳动物细胞中产生的市售rhuEPO来制造脱唾液酸rhuEPO。基于现有的知识和技术，很可能对植物进行遗传修饰，以生产不含唾液酸的人类糖蛋白。我们在烟草中共表达了人GalT基因和与ER信号肽融合的EPO基因，并尝试以低成本生产具有大规模生产潜力的去唾液酸rhuEPO。我们预期植物产生的去唾液酸rhuEPO将具有与酶促产生的去唾液酸rhuEPO类似的细胞保护功能。本项目的结果也将为植物糖基化机制和植物产生的无唾液酸rhuEPO的抗凋亡机制提供见解。此外，所建立的蛋白质纯化方法和糖蛋白分析方法将对未来的植物糖工程项目非常有价值。公共卫生相关性：我们的目标是转基因植物生产脱唾液酸rhuEPO-一种特殊的人类蛋白质。这种类型的生物药物蛋白可以作为神经治疗和心脏保护剂进行开发。利用植物表达系统，我们期望生产一种廉价的和功能性的去唾液酸rhuEPO，以防止细胞死亡与大规模生产的潜力。