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Basic study on mass production of therapeutic proteins for serious disease by genetically engineered rice cell

利用基因工程水稻细胞大规模生产严重疾病治疗蛋白的基础研究

基本信息

批准号：
11650821
负责人：
TERASHIMA Masaaki
金额：
$ 2.3万
依托单位：
Osaka Prefecture University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

项目摘要

The following significant results were obtained from the research on production of therapeutic protein, α_1-antitrypsin (AAT) by genetically engineered rice cell.(1) Growth rate of genetically engineered cell was the same as that of non-transformed rice cell. Specific growth rate of rice cell, 0.3 d^<-1>, was of the same order of magnitude as those of other plant cells.(2) Oxygen transfer coefficient K_La was 7-8 h^<-1>, and aeration rate did not affect the growth rate of rice cell.(3) Ammonium was produced at cell growth phase and AAT production phase. However, ammonium did not affect the AAT productivity.(4) Adjustment of osmotic pressure between growth medium and production medium has favorable effective for AAT production.(5) AAT productivity was tripled using pyrvic acid as an altemative carbon source.(6) Continuous production was achieved in the continuous stirred tank reactor.(7) Cell density was increased up to 14 g-dry cell/L in the batch-type reactor.(8) Since maximum productivity of AAT at optimal condition was 25 mg/g-dry cell, at least 350 mg/L of AAT can be produced in batch-type reactor.The recombinant proteins can be rapidly induced by sugar depletion in the employed expression system. Our research strongly suggests that protein production system more efficient than microbial system can be constructed by the rice cell culture system. We hope the mass production system using plant cell culture can be achieved by exploring process-engineering research based on this fundamental research.

利用水稻基因工程细胞生产治疗性蛋白α_1-抗胰蛋白酶（AAT）的研究获得了以下重要结果：(1)转基因细胞的生长速度与未转化水稻细胞相同。水稻细胞的比生长率为0.3 d^<-1>，与其他植物细胞的比生长率处于同一数量级。(2)氧传递系数K_La为7 ~ 8 h^<-1>，曝气速率不影响水稻细胞的生长速率。(3)铵在细胞生长期和AAT产生期产生。铵对AAT的产率没有影响。(4)生长介质与生产介质间渗透压的调节对AAT的生产有良好的效果。(5)以丙酮酸为碳源，AAT产率提高3倍。(6)在连续搅拌槽式反应器中实现了连续生产。(7)间歇式反应器中细胞密度提高到14 g-dry Cell /L。(8)由于AAT在最佳条件下的最大产率为25 mg/g-干电池，因此间歇式反应器至少可生产350 mg/L的AAT。在所采用的表达体系中，可以通过缺糖快速诱导重组蛋白。我们的研究强烈表明，水稻细胞培养系统可以构建比微生物系统更高效的蛋白质生产系统。我们希望在此基础研究的基础上，通过探索工艺工程研究，实现植物细胞培养的批量生产系统。