Metabolic Engineering of Synechocystis sp Strain PCC 6803 for Isobutanol Production
Metabolic Engineering of Synechocystis sp Strain PCC 6803 for Isobutanol Production
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DOI：
10.1128/aem.02827-12
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发表时间：
2013-02-01
影响因子：
4.4
通讯作者：
Tang, Yinjie J.
中科院分区：
文献类型：
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作者：
Varman, Arul M.;Xiao, Yi;Tang, Yinjie J.
全球变暖和化石燃料储量的减少促使人们对从可再生资源合成先进生物燃料产生了极大的兴趣。为了解决这些问题，我们对蓝细菌集胞藻属菌株PCC 6803进行了代谢工程改造，以开发一种可以在自养和兼养条件下合成异丁醇的菌株。在表达来自埃利希途径的两个异源基因的情况下，工程菌株可以在气密摇瓶中从50 mM碳酸氢盐积累90 mg/L的异丁醇。该菌株不需要任何诱导物（即，异丙基β-D-1-硫代吡喃半乳糖苷[IPTG]）或抗生素以维持其异丁醇产量。在葡萄糖的存在下，异丁醇的合成仅被适度地促进（滴度= 114 mg/升）。同位素分析结果表明，与野生型菌株相比，突变株的葡萄糖利用率显著降低，主要利用自养代谢进行生物量的增长和异丁醇的生产。由于异丁醇是有毒的细胞，也可能被光化学降解的羟基自由基在培养过程中，我们采用了原位去除的异丁醇使用油醇作为溶剂陷阱。这导致在兼养培养条件下异丁醇的最终净浓度为298 mg/升。
Global warming and decreasing fossil fuel reserves have prompted great interest in the synthesis of advanced biofuels from renewable resources. In an effort to address these concerns, we performed metabolic engineering of the cyanobacterium Synechocystis sp. strain PCC 6803 to develop a strain that can synthesize isobutanol under both autotrophic and mixotrophic conditions. With the expression of two heterologous genes from the Ehrlich pathway, the engineered strain can accumulate 90 mg/liter of isobutanol from 50 mM bicarbonate in a gas-tight shaking flask. The strain does not require any inducer (i.e., isopropyl beta-D-1-thiogalactopyranoside [IPTG]) or antibiotics to maintain its isobutanol production. In the presence of glucose, isobutanol synthesis is only moderately promoted (titer = 114 mg/liter). Based on isotopomer analysis, we found that, compared to the wild-type strain, the mutant significantly reduced its glucose utilization and mainly employed autotrophic metabolism for biomass growth and isobutanol production. Since isobutanol is toxic to the cells and may also be degraded photochemically by hydroxyl radicals during the cultivation process, we employed in situ removal of the isobutanol using oleyl alcohol as a solvent trap. This resulted in a final net concentration of 298 mg/liter of isobutanol under mixotrophic culture conditions.