METABOLIC ENGINEERING OF CLOSTRIDIUM-ACETOBUTYLICUM ATCC-824 FOR INCREASED SOLVENT PRODUCTION BY ENHANCEMENT OF ACETONE FORMATION ENZYME-ACTIVITIES USING A SYNTHETIC ACETONE OPERON
METABOLIC ENGINEERING OF CLOSTRIDIUM-ACETOBUTYLICUM ATCC-824 FOR INCREASED SOLVENT PRODUCTION BY ENHANCEMENT OF ACETONE FORMATION ENZYME-ACTIVITIES USING A SYNTHETIC ACETONE OPERON
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DOI：
10.1002/bit.260420906
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发表时间：
1993-11-05
影响因子：
3.8
通讯作者：
BENNETT, GN
中科院分区：
文献类型：
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作者：
MERMELSTEIN, LD;PAPOUTSAKIS, ET;BENNETT, GN
通过基因改变乙酰丁基梭菌的产物形成能力对于通过发酵继续工业化生产丁醇和丙酮是必要的。利用乙酰丁酸杆菌ATCC 824(PFNK6)在pH 4.5、5.5或6.5的条件下进行分批发酵。PFNK6含有一个合成操纵子，其中三个同源丙酮形成基因(ADC、CTFA和ctfB)是从ADC启动子转录而来的。相应的酶(乙酰乙酸脱羧酶和辅酶A-转移酶)在pH为4.5的发酵条件下表达效果最好。然而，在pH值为5.5时，溶剂含量最高。在pH值为5.5的条件下，ATCC 824(PFNK6)的丙酮、丁醇和乙醇的最终浓度分别比无质粒对照菌株高95%、37%和90%；葡萄糖的溶剂产率(g/g)高50%；残留羧酸的质量降低22倍。在所有pH值下，ATCC 824(PFNK6)的丙酮形成酶比对照发酵更早表达，从而导致丙酮形成的更早诱导。此外，菌株ATCC 824(PFNK6)在发酵过程中显著提前产生丁醇，并在pH 6.5时产生大量溶剂。菌株ATCC 824在pH 6.5时只产生微量的溶剂。与ATCC 824相比，含有不含Ace操纵子的载体的质粒对照菌株也产生更高水平的溶剂[尽管低于菌株ATCC 824(PFNK6)]和更低水平的酸。含有Ace操纵子的质粒携带的衍生物的菌株，其中单独表达乙酰乙酸脱羧酶或CoA转移酶的水平较高，产生的酸和溶剂的水平与质粒对照菌株相似。(C)1993年John Wiley&Sons，Inc.
The ability to genetically alter the product-formation capabilities of Clostridium acetobutylicum is necessary for continued progress toward industrial production of the solvents butanol and acetone by fermentation. Batch fermentations at pH 4.5, 5.5, or 6.5 were conducted using C. acetobutylicum ATCC 824 (pFNK6). Plasmid pFNK6 contains a synthetic operon (the ''ace operon'') in which the three homologous acetone-formation genes (adc, ctfA, and ctfB) are transcribed from the adc promoter. The corresponding enzymes (acetoacetate decarboxylase and CoA-transferase) were best expressed in pH 4.5 fermentations. However, the highest levels of solvents were attained at pH 5.5. Relative to the plasmid-free control strain at pH 5.5, ATCC 824 (pFNK6) produced 95%, 37%, and 90% higher final concentrations of acetone, butanol, and ethanol, respectively; a 50% higher yield (g/g) of solvents on glucose; and a 22-fold lower mass of residual carboxylic acids. At all pH values, the acetone-formation enzymes were expressed earlier with ATCC 824 (pFNK6) than in control fermentations, leading to earlier induction of acetone formation. Furthermore, strain ATCC 824 (pFNK6) produced butanol significantly earlier in the fermentation and produced significant levels of solvents at pH 6.5. Only trace levels of solvents were produced by strain ATCC 824 at pH 6.5. Compared with ATCC 824, a plasmid-control strain containing a vector without the ace operon also produced higher levels of solvents [although lower than those of strain ATCC 824 (pFNK6)] and lower levels of acids. Strains containing plasmid-borne derivatives of the ace operon, in which either the acetoacetate decarboxylase or CoA-transferase alone were expressed at elevated levels, produced acids and solvents at levels similar to those of the plasmid-control strain. (C) 1993 John Wiley & Sons, Inc.