Engineering a Metabolic Pathway for Isobutanol Biosynthesis in Bacillus subtilis
Engineering a Metabolic Pathway for Isobutanol Biosynthesis in Bacillus subtilis
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枯草芽孢杆菌异丁醇生物合成代谢途径的工程设计
DOI：
10.1007/s12010-011-9268-1
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发表时间：
2012-09-01
影响因子：
3
通讯作者：
Wen, Jianping
中科院分区：
文献类型：
--
作者：
Jia, Xiaoqiang;Li, Shanshan;Wen, Jianping
异丙酮酸脱羧酶(KDC)和乙醇脱氢酶(ADH)催化α-酮基异戊酸酯生物合成异丁醇。在这项工作中，异丁醇的生物合成途径在枯草芽孢杆菌中设计，枯草芽孢杆菌是一种显著的耐溶剂宿主。为此，构建了枯草杆菌强启动子P-43控制下的KDC和ADH表达载体pPKA。携带pPKA基因的重组枯草杆菌的产物中检测到异丁醇，而野生型菌株未检测到异丁醇。考察了葡萄糖浓度、缬氨酸投加量等培养基组分和初始pH、接种量、装液量等操作参数对异丁醇产量的影响。在葡萄糖浓度为3%，缬氨酸添加量为2%，初始pH为7.0，接种量为1%，装液量为50mL/250mL的条件下，发酵35h，异丁醇产量达到最高值0.607 g/L。虽然异丁醇产量较低，但这是首次在工程菌中成功生产异丁醇的尝试，显示了其作为异丁醇生产细胞工厂的巨大潜力。
Isobutanol can be biosynthesized via alpha-ketoisovalerate catalyzed by heterologous keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH). In this work, isobutanol biosynthesis pathway was designed in Bacillus subtilis, a notable solvent-tolerant host. In order to do that, a plasmid pPKA expressing KDC and ADH under the control of a B. subtilis strong promoter P-43 was constructed. Isobutanol was detected in the products of the recombinant B. subtilis harboring pPKA plasmid, whereas none was detected by the wild-type strain. Effects of the medium ingredients such as glucose concentration and valine addition, and operating parameters such as initial pH, inoculation volume, and medium work volume on isobutanol production were also investigated. Isobutanol production reached to the maximum of 0.607 g/L after 35-h cultivation under the conditions: glucose concentration of 3%, valine addition of 2%, initial pH of 7.0, inoculum of 1%, and work volume of 50 mL/250 mL. Though the isobutanol production by the recombinant was low, it was the first successful attempt to produce isobutanol in engineered B. subtilis, and the results showed its great potential as an isobutanol-producing cell factory.