ENHANCEMENT OF ACTIVITY OF MICROORGANISMS CULTIVATED WITH BIOMASS SUPPORT PARTICLES

增强用生物质载体颗粒培养的微生物的活性

• 批准号: 04650872
• 负责人: ISHIKAWA Haruo
• 金额: $ 1.28万
• 依托单位: Osaka Prefecture University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04650872/
• 关键词: Porous urethane beads; Immobilized microorganisms; Filamentous fungus; Rhizopus chinensis; Lipase; Enzyme purification; Ester-hydrolyzing reaction; Interesterification reaction; 固定化培養; 固定化微生物菌体

This work was undertaken to develop a new method for producing immobilized microorganisms which have high activity and are suitable for industrial use. First, proteins in the cells and supernatant, which were produced by suspension culture of filamentous fungus Rhizopus chinensis and by cultivating the cells in porous urethane beads, were analyzed by using SDS-PAGE.Then, one of the proteins in supernatant obtained by suspension culture of Rh. chinensis was purified. The principla results are as follows : (1) The SDS-PAGE of the proteins in the cells and in the supernatant, both of which were obtained by suspension culture and by cultivating the cells in porous urethane beads, showed that the patterns of electrophoresis were considerably different. This means that different kinds of proteins were produced in the supernatant and cells, and this depended on the cultivation methods. (2) One of the proteins in the supernatant of the suspension culture was purified by use of ion-exchange chromatography, hydrophobic interaction chromatography and gel filtration chromatography. (3) The purified protein was a monomeric enzyme consisting of a single polypeptide chain and had a molecular weight of 32,000. The optimum reaction temperature and optimum pH were 37ﾟC and 5.5, respectively. A lyophilized sample of the purified protein had an ester-hydrolyzing activity of 81.8 Unit/mg, but did not have any interesterification activity. From these results, we concluded that Rh. chinensis secreted lipase having high ester-hydrolyzing activity and have lipase with high interesterification activity in the cells.

这项工作是为了开发一种新的生产固定化微生物的方法，该固定化微生物具有高活性，适合工业使用。首先，对丝状真菌中国根霉悬浮培养产生的细胞和细胞培养上清液中的蛋白质进行了SDS-PAGE分析，得到了Rh悬浮培养上清液中的蛋白质。得到了纯化的中华草。主要结果如下：(1)细胞悬浮培养和细胞培养上清液中蛋白质的SDS-PAGE结果显示，细胞内蛋白质和细胞培养上清液中蛋白质的电泳图谱有较大差异。这意味着在培养上清和细胞中产生了不同种类的蛋白质，这取决于培养方法。(2)用离子交换层析、疏水作用层析和凝胶过滤层析对悬浮培养上清液中的一种蛋白质进行了纯化。(3)纯化的蛋白质是由单一多肽链组成的单体酶，相对分子质量为32,000。最适反应温度为37ﾟC，最适pH为5.5。纯化蛋白的冻干样品具有81.8U/mg的酯水解活性，但没有任何酯交换活性。根据这些结果，我们得出结论：Rh.中国对虾分泌的脂肪酶具有较高的酯水解酶活性，细胞内存在具有较高酯交换活性的脂肪酶。