[1]彭 惠,毛忠贵,武国干,等.嗜热厌氧乙醇菌JW200中乙醛脱氢酶的纯化[J].南京师大学报(自然科学版),2007,30(01):78-81.
　Peng Hui,Mao Zhonggui,Wu Guogan,et al.Purification of Acetaldehyde Dehydrogenase From Thermoanaerobacter ethanolicus JW200[J].Journal of Nanjing Normal University(Natural Science Edition),2007,30(01):78-81.
点击复制

嗜热厌氧乙醇菌JW200中乙醛脱氢酶的纯化()
分享到：

《南京师大学报（自然科学版）》[ISSN:1001-4616/CN:32-1239/N]

卷:
第30卷
期数:
2007年01期
页码:
78-81
栏目:
生命科学
出版日期:
2007-03-30

文章信息/Info

Title:
Purification of Acetaldehyde Dehydrogenase From Thermoanaerobacter ethanolicus JW200
作者:
彭  惠1;  毛忠贵1;  武国干2;  邵蔚蓝1;  2
( 1. 江南大学工业生物技术教育部重点实验室, 江苏无锡214036 )
( 2. 南京师范大学微生物工程重点实验室, 江苏南京210097 )
Author(s):
Peng Hui1; Mao Zhonggui1; Wu Guogan2; Shao Weilan1; 2*
1.The Key Laboratory of Industrial Biotechnology Under Ministry of Education,Southern Yangtze University,Wuxi 214036,China
2. The Key Laboratory ofM icrob iolE ngineering, Nan jing Norm alUn ivers ity, Nan j ing 210097, Ch in a
关键词:
乙醛脱氢酶;  嗜热厌氧乙醇菌;  纯化
Keywords:
ace taldehyde dehydrogenase;  Thermoanaerobacter ethano licus;  pur ification
分类号:
Q814.1
摘要:
研究了嗜热厌氧乙醇菌(Thermoanaerobacter ethanolicus)JW200中乙醇代谢途径的关键酶之一乙酰CoA依赖型的乙醛脱氢酶(acetaldehyde dehydrogenase,ALDH,EC 1.2.1.10)的纯化.结果表明,经DEAE Sepha-rose Fast Flow阴离子交换层析、两次为30%与80%硫酸铵盐析和相应盐浓度的Butyl-HIC疏水层析、TOYOPE-ARL HW-55F分子筛层析等提纯步骤,可得到电泳纯的ALDH,其提纯倍数为910倍,得率为7%.由SDS-PAGE和梯度PAGE测得全酶由4个亚基组成,全酶相对分子质量为360 000,亚基相对分子质量为100 000.
Abstract:
A aceta ldehyde dehydrogenase ( CoA-acety la ting) w hich is one o f the key enzym es o f the a lco- ho lm etabo lic pathw ay w as pur ified from Thermoanaerobacter ethano licus JW 200 through follow ing steps: ① ion exchang e chrom atography on DEAE-Sepharose Fast Flow, ② amm on ium su lfate fractionation w ith saturation 30% and hydrophob ic in teraction chrom atography on Butyl- H IC, ③ amm onium su lfate frac tionation w ith saturation 80% and hydrophob ic interac tion chrom atography on Bu ty l-H IC, ④ ge l filtration on TOYOPEARL HW - 55F. The pur ified enzyme showed a sing le band on SDS-po lyac ry lam ide ge l e lec tropho resis w ith a pu rifica tion o f 910 fo ld, and a y ie ld o f 7%. The mo lecu la rw e ight o f the subun it and the w ho le enzyme w ere estim ated by SDS-PAGE and g radation PAGE as 100 000 and 360 000, respective ly.

参考文献/References:

[ 1] Y Runtao, Chen J S. Coenzym e A-acy la ting a ldehyde dehydrogenase from C lostr id ium be ijerinckii NRRL B592[ J] . Appl
Env irM ic ro, 1990, 56( 9): 2 591-2 599.
[ 2] W ieg el J. Therm oanaerobacte r e thano licus g en. nov. , spec. nov. , a new, ex trem e thermophilic, anaerobic bacter ium [ J]. Arch M icrob io ,l 1981, 128( 5): 343-348.
[ 3] H ildH M, Stuckey D C, Leak D J. Effect of nutr ient lim itation on product forma tion dur ing continuous ferm en tation o f xylose with Therm oanaerobacter ethano licus JW200 Fe( 7) [ J]. ApplM ic robiol B iotechno,l 2003, 60( 12): 679-686.
[ 4] Burde tte D S, V ieille C, Zeikus J G. C loning and expression of the gene encoding the Therm oanaerobacter ethano licus 39E secondary-alcoho l dehydrogenase and b iochem ica l charac terization o f the enzym e[ J]. B iochem J, 1996, 316( 1) : 115-122.
[ 5] Burdette D S, Ze ikus J G. Purification of aceta ldehyde dehydrogenase and alcoho l dehy-drogenases from Therm oanaerobacter ethanolicus 39E and charac terization o f the se-condary alcoho l dehydrogenase ( 2A` dh) as a bifunctiona l alcoho l dehydrogenase-acetyl-CoA reductive th ioesterase[ J]. B io chem J, 1994, 302( 3): 163-170.
[ 6] 汪家政, 范明. 蛋白质技术手册[M ]. 北京: 科学出版社, 2001: 42-122.
[ 7] Sanche z L B. A ldehyde dehydrogenase ( CoA-acetylating ) and the m echan ism of etha-no l form ation in the am ito chondriate protist, G iard ia lam blia[ J]. Arch ives B iochem istry and B iophysics, 1998, 353( 1): 57-64.
[ 8] To th J, Chen J S. The a ld gene, encoding a coenzym eA-Acy la ting a ldehyde de-hydrogenase, distingu ishes c lostr id ium beijerinck ii and two o ther so lvent producing clos-tr idia from c lo strid ium ace tobu ty llicum [ J]. App l Env irM icro, 1999, 65 ( 11):4 973-4 980.

备注/Memo

备注/Memo:
基金项目: 中国973 项目子课题基金( 2004CB719600)、国家自然科学基金( 30170511 )资助项目.
作者简介: 彭  惠( 1979- ) , 女, 博士研究生, 主要从事极端微生物遗传代谢工程的学习与研究. E-m ail:pph0259@ 126. com
通讯联系人: 邵蔚蓝( 1958-) , 女, 特聘教授, 博士生导师, 主要从事微生物分子生物学的教学与研究. E-m ail:w lshao@ jsm ail. com. cn
更新日期/Last Update: 2013-05-05