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利用新型 Corynebacterium crenatum 表达系统表达玻璃珠菌血红蛋白增强 L-精氨酸的生产。

Enhanced production of L-arginine by expression of Vitreoscilla hemoglobin using a novel expression system in Corynebacterium crenatum.

机构信息

The Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, People's Republic of China.

出版信息

Appl Biochem Biotechnol. 2011 Mar;163(6):707-19. doi: 10.1007/s12010-010-9076-z. Epub 2010 Sep 11.

Abstract

Corynebacterium crenatum SYPA 5-5 is an aerobic and industrial L: -arginine producer. It was proved that the Corynebacterium glutamicum/Escherichia coli shuttle vector pJC1 could be extended in C. crenatum efficiently when using the chloramphenicol acetyltransferase gene (cat) as a reporter under the control of promoter tac. The expression system was applied to over-express the gene vgb coding Vitreoscilla hemoglobin (VHb) to further increase the dissolved oxygen in C. crenatum. As a result, the recombinant C. crenatum containing the pJC-tac-vgb plasmid expressed VHb at a level of 3.4 nmol g(-1), and the oxygen uptake rates reached 0.25 mg A(562)(-1)  h(-1) which enhanced 38.8% compared to the wild-type strain. Thus, the final L: -arginine concentration of the batch fermentation reached a high level of 35.9 g L(-1), and the biomass was largely increased to 6.45 g L(-1), which were 17.3% and 10.5% higher than those obtained by the wild-type strain, respectively. To our knowledge, this is the first report that the efficient expression system was constructed to introduce vgb gene increasing the oxygen and energy supply for L: -arginine production in C. crenatum, which supplies a good strategy for the improvement of amino acid products.

摘要

棒状杆菌 SYPA5-5 是一株需氧、生产 L-精氨酸的工业菌株。实验证明,在协同利用氯霉素乙酰转移酶基因(cat)作为报告基因,受 tac 启动子调控的情况下,谷氨酸棒杆菌/大肠杆菌穿梭载体 pJC1 可有效地在棒状杆菌中进行扩展。利用该表达系统过表达编码黏细菌血红蛋白(VHb)的基因 vgb,进一步提高了棒状杆菌中的溶解氧。结果,含有 pJC-tac-vgb 质粒的重组棒状杆菌表达 VHb 的水平达到 3.4 nmol/g,比野生型菌株提高了 38.8%,氧摄取率达到 0.25 mg A(562)(-1)h(-1)。因此,分批发酵的最终 L-精氨酸浓度达到了 35.9 g/L,生物量也大大增加到 6.45 g/L,分别比野生型菌株提高了 17.3%和 10.5%。据我们所知,这是首次构建高效表达系统,引入 vgb 基因提高棒状杆菌中 L-精氨酸生产的氧气和能量供应,为氨基酸产品的改良提供了一个良好的策略。

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