Please wait a minute...

中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志  2010, Vol. 30 Issue (08): 93-99    DOI: Q785 Q786
技术与方法     
一步法生产1,5-戊二胺谷氨酸棒杆菌基因工程菌的构建
牛涛,黎明,张俊环,宋馨宇,张建中
工业微生物教育部重点实验室天津科技大学 生物工程学院 天津 300457
Construction of Recombinant Corynebacterium glutamicum Producing 1,5-Pentanediamine by One Step Method
NIU Tao,LI Ming,ZHANG Jun-huan,SONG Xin-yu,ZHANG Jian-zhong
Key Laboratory of Industrial Microbiology, Ministry of Education, The College of Bioengineering, Tianjin University of Science and Technology, Tianjin 300457, China
 全文: PDF(562 KB)   HTML
摘要:

1,5-戊二胺是一种重要的化工原料,发酵法生产1,5-戊二胺是一条新颖且具有潜在竞争力的生产途径。以蜂房哈夫尼菌(Hafnia alvei)AS1.1009基因组为模板,通过PCR扩增,得到大小约为2.2 kb的赖氨酸脱羧酶基因ldc。以大肠杆菌(Escherichia coli) /谷氨酸棒杆菌(Corynebacterium glutamicum)穿梭质粒pXMJl9为载体,将扩增得到的目的基因片段克隆至谷氨酸棒杆菌C. glutamicum TK260512,获得重组菌株C. glutamicum TK260512/pXMJl9-ldc.在摇瓶发酵水平上,通过IPTG诱导ldc基因的表达,并采用反相高效液相色谱方法测定了发酵液中1,5-戊二胺的含量,结果显示,经36 h发酵,工程菌C. glutamicum TK260512/pXMJ19-ldc的1,5-戊二胺产量为0.96 g/L。

关键词: 赖氨酸脱羧酶谷氨酸棒杆菌1,5-戊二胺    
Abstract:

1,5-Pentanediamine is one of the most important industrial chemicals for its highly desired properties and its wide applications as a key component of an emerging polymer business. Biological production of 1,5-Pentanediamine has been a novel and competitive way. An about 2.2 kb fragment of Hafnia alvei lysine decarboxylase gene encoding lysine decarboxylase (LDC) was proliferated by polymerase chain reaction by using chromosomal DNA of Hafnia alvei as the template. The obtained ldc fragment was inserted into E. coli/C. glutamicum shuttle vector pXMJ19 to construct an expression plasmid pXMJ19-ldc. It was then introduced into C. glutamicum TK260512 via electrotransformation, and a recombinant C. glutamicum TK260512/pXMJl9-ldc was obtained. The 1,5-Pentanediamine in the broth was detected by HPLC when the culture in the shake flask was indueed with 0.5 mmol/L IPTG. The results showed that after 36 h cultivation,the recombinant stain could produce 0.96 g/L 1,5-Pentanediamine.

Key words: Lysine decarboxylase    C.glutamicum    1,5-pentanediamin
收稿日期: 2010-03-24 出版日期: 2010-08-25
通讯作者: 黎明     E-mail: liming09@tust.edu.cn
服务  
把本文推荐给朋友
加入引用管理器
E-mail Alert
RSS
作者相关文章  
牛涛
黎明
张俊环
宋馨宇
张建中

引用本文:

牛涛 黎明 张俊环 宋馨宇 张建中. 一步法生产1,5-戊二胺谷氨酸棒杆菌基因工程菌的构建[J]. 中国生物工程杂志, 2010, 30(08): 93-99.

NIU Chao, LI Meng, ZHANG Dun-Huan, SONG Xin-Yu, ZHANG Jian-Zhong. Construction of Recombinant Corynebacterium glutamicum Producing 1,5-Pentanediamine by One Step Method. China Biotechnology, 2010, 30(08): 93-99.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/Q785 Q786        https://manu60.magtech.com.cn/biotech/CN/Y2010/V30/I08/93

[1] 陈学好, 于杰, 李伶利. 高等植物开花结实的多胺研究进展. 植物学通报, 2003, 20(1): 3642. Chen X H, Yu J, Li L L. Chinese Bulletin of Botany, 2003,20(1): 3642. 
[2] 王晓云, 邹琦. 多胺与植物衰老关系研究进展. 植物学通报, 2002, 19(1): 1120. Wang X Y, Zou Q, Chinese Bulletin of Botany, 2002,19(1): 1120. 
[3] Casalino M, Latella M C, Prosseda G, et al. Molecular evolution of the lysine decarboxylasedefective phenotype in Shigella sonnei. International Journal of Medical Microbiology. 2005, 294: 503512. 
[4] Kiyohiko N, Shuichi E, Yukiko M. Enzymatic method for producing cadaverine dicarboxylate and its use for the production of nylon.Japan, 147 688, 20030526. 
[5] Francisco J, Flores J, Rincón J F. Characterization of the ironregulated desA promoter of Streptomyces pilosus as a system for controlled gene expression in actinomycetes. Microb Cell Fact. 2003, 71: 339349. 
[6] Delavega A L, Delcour A H. Cadaverine induces closing of E.coli porins. EMBO, 1995, 14(23): 60586065. 
[7] Hirao T, Sato M, Shirahata A. Covalent linkage of polyamines to peptidoglycan in Anaerovibrio lipolytica. Bacteriol, 2000, 182(4): 11541157. 
[8] Kamio Y, Itoh Y, Terawaki Y. Chemical structure of peptidoglycan in Selenomonas ruminantium: cadaverine links covalently to the Dglutamic acid residue of peptidoglycan. Bacteriol, 1981, 146(1): 4953. 
[9] Kamio Y, Nakamura K. Putrescine and cadaverine are constituents of peptidoglycan in Veillonella alcalescens and Veillonella parvula. Bacteriol,1987, 169(6): 28812884. 
[10] Takashi M, Hideki S, Masahiro H, et al. Metabolic engineering of Corynebacterium glutamicum for cadaverine fermentation. Biosci Biotechnol Biochem, 2007, 71(9):21302135. 
[11] Tateno T, Okada Y, Tsuchidate T, et al. Direct production of cadaverine from soluble starch using Corynebacterium glutamicum coexpressing αamylase and lysine decarboxylase. Appl Microbiol Biotechnol, 2009, 82:115121. 
[12] 蒋丽丽, 刘均忠, 沈俞,等. 用固定化赖氨酸脱羧酶细胞制备1,5戊二胺. 精细化工, 2007, 24(11):10801084. Jiang L L, Liu J Z, Shen Y, et al. Fine Chem Icals, 2007, 24(11): 10801084. 
[13] Tateno T, Fukuda H, Kondo A. Production of Llysine from starch by Corynebacterium glutamicum displaying αamylase on its cell surface. Appl Microbiol Biotechnol, 2007, 74:1213–1220. 
[14] 吴健胜. 赖氨酸脱羧酶对水稻白叶枯病菌有机酸毒素的解毒作用及其基因的克隆和表达研究及草酸解毒菌的初步研究. 南京: 南京农业大学, 2000. Wu J S. Cloning and expression of the gene encoding a lysine decaroxylase and activity of the enzyme in detoxifying organic acid toxin produced by xanthomonas oryzae pv. oryzae and a preliminary study on detoxification of oxalate toxin by a bacterial strain. Nanjing:Nanjing Agricultural University ,2000. 
[15] Sambrook J, Fritsch E F, Maniatis T. Molecular Cloning: A Laboratory Manual. 2nd ed. New York: Cold Spring Harbor Laboratory Press, 1989. 
[16] Santamaria R, Gil J A, Mesas J M, et al. Characterization of an endogenous plasmid and development of cloning vectors and a transformation system in Breuibacterium lactofermentum. Journal of General Microbiology, 1984,130: 22372246. 
[17] Amador E, Castro J M, Correia A, et al.Structure and organization of the rrnD operon of Brevibacterium lactofermentum: analysis of the 16S rRNA gene. Microbiol, 1999, 145:915924. 
[18] van der Rest M E, Lange C, Molenaar D. A heat shock following electroporation induces highly efficient transformation of Corynebacterium glutamicum with xenogenetic plasmid DNA . Appl Microbio Biotech, 1999, 52: 541545. 
[19] 赵传孝, 韩绍英. 食品检验技术手册. 中国食品出版社, 1990.696697. Zhao C X, Han S Y. Technical Manual of Food Analysis. Food Publishing Company of China, 1990. 696697. 
[20] Tateno T, Fukuda H, Kondo A. Production of Llysine from starch by Corynebacterium glutamicum displaying αamylase on its cell surface. Appl Microbiol Biotechnol, 2007, 74: 12131220. 
[21] Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of proteindye binding. Anal Biochem ,1976,72:248254. 
[22] Donna L S, Elizabeth A B, Breck B, et al. Purification and physical propertieson inducible Escherichia coli lysine decarboxylase. Biochemistry, 1974, 13: 662676.

[1] 万方, 陈民良, 张斌, 陈进聪, 陈雪岚. 代谢工程改造微生物高产氨基酸的策略[J]. 中国生物工程杂志, 2015, 35(3): 99-103.
[2] 叶菁, 许敬亮, 肖波, 袁振宏, 徐惠娟, 杨柳, 李谢昆. 谷氨酸棒杆菌戊糖代谢利用研究进展[J]. 中国生物工程杂志, 2012, 32(11): 132-136.
[3] 叶菁, 许敬亮, 肖波, 袁振宏, 徐惠娟, 杨柳, 李谢昆. 谷氨酸棒杆菌戊糖代谢利用研究进展[J]. 中国生物工程杂志, 2012, 32(11): 132-136.
[4] 金雪霞, 张晓梅, 窦文芳, 许泓瑜, 许正宏. 谷氨酸棒杆菌SYPS-062 L-丝氨酸脱水酶活性分析及其基因敲除对L-丝氨酸积累的影响[J]. 中国生物工程杂志, 2011, 31(01): 29-34.
[5] 张博, 李铁民, 杨智勇, 胡永飞, 李玉. 谷氨酸棒杆菌H+-ATPase基因失活提高谷氨酸产生量[J]. 中国生物工程杂志, 2011, 31(01): 35-39.
[6] 任建洪,张晓梅,窦文芳,许泓瑜,许正宏. 不同来源的谷氨酸棒杆菌氨基脱氧分支酸合成酶的活性分析[J]. 中国生物工程杂志, 2009, 29(08): 57-61.
[7] 张晓娟,窦文芳,许泓瑜,许正宏. 维生素对谷氨酸棒杆菌SYPS-062直接发酵合成L-丝氨酸的影响[J]. 中国生物工程杂志, 2007, 27(5): 50-55.
[8] 余秉琦, 沈微, 诸葛健. 适用于异源DNA高效整合转化的谷氨酸棒杆菌电转化法[J]. 中国生物工程杂志, 2005, 25(02): 78-81.