Engineering the genotype of Acinetobacter sp strain ADP1 to enhance biosynthesis of cyanophycin
Engineering the genotype of Acinetobacter sp strain ADP1 to enhance biosynthesis of cyanophycin
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DOI：
10.1128/aem.72.2.1410-1419.2006
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发表时间：
2006-02-01
影响因子：
4.4
通讯作者：
Steinbüchel, A
中科院分区：
文献类型：
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作者：
Elbahloul, Y;Steinbüchel, A
为了研究精氨酸供应和磷酸限制对不动杆菌ADPI中蓝藻素（CGP）合成和积累的重要性，失活了编码精氨酸调节蛋白（argR）和精氨酸琥珀酰转移酶（ast，4）的基因，并分析了这些突变对CGP合成的影响。当细胞在谷氨酸盐上生长时，这些基因的失活分别导致CGP含量增加3.5倍或7倍。两个基因中的敲除突变导致更好地理解在不动杆菌属菌株ADPI的细胞生长期间向精氨酸添加其他底物对CGP合成的影响。如果使用氨基酸作为碳源，则ArgF（鸟氨酸氨甲酰转移酶）、CarA-CarB（氨甲酰磷酸合成酶的小亚基和大亚基）和PepC（磷酸烯醇丙酮酸羧化酶）的过表达触发CGP的合成，而葡萄糖酸盐或其他糖不触发CGP的合成。不动杆菌属菌株ADPI的细胞，这是在很大程度上缺乏碳水化合物代谢的基因，显示出CGP含量显着增加时，生长在矿物培养基中补充谷氨酸，天冬氨酸，或精氨酸。不动杆菌Δ astA（pYargF）菌株不能利用精氨酸，但合成更多的精氨酸，导致CGP含量高达30%和25%的细胞干物质时，生长在protamylasse或Luria-Bertani培养基，分别。这种重组菌株克服了昂贵的精氨酸供应的瓶颈，其中它产生约75%的从在含有纯精氨酸作为唯一碳源的矿物培养基上生长的亲本细胞获得的CGP。磷酸盐饥饿是唯一已知的触发CGP合成在这种细菌，它拥有的PhoB/PhoR磷酸调节子系统。phoB的过表达导致CGP含量增加8.6倍，在一个非限制性的磷酸盐浓度的亲本菌株相比。
To study the importance of arginine provision and phosphate limitation for synthesis and accumulation of cyanophycin (CGP) in Acinetobacter sp. strain ADPI, genes encoding the putative arginine regulatory protein (argR) and the arginine succinyltransferase (ast,4) were inactivated, and the effects of these mutations on CGP synthesis were analyzed. The inactivation of these genes resulted in a 3.5- or 7-fold increase in CGP content, respectively, when the cells were grown on glutamate. Knockout mutations in both genes led to a better understanding of the effect of the addition of other substrates to arginine on CGP synthesis during growth of the cells of Acinetobacter sp. strain ADPI. Overexpression of ArgF (ornithine carbamoyltransferase), CarA-CarB (small and large subunits of carbamoylphosphate synthetase), and PepC (phosphoenolpyruvate carboxylase) triggered synthesis of CGP if amino acids were used as a carbon source whereas it was not triggered by gluconate or other sugars. Cells of Acinetobacter sp. strain ADPI, which is largely lacking genes for carbohydrate metabolism, showed a significant increase in CGP contents when grown on mineral medium supplemented with glutamate, aspartate, or arginine. The Acinetobacter sp. Delta astA(pYargF) strain is unable to utilize arginine but synthesizes more arginine, resulting in CGP contents as high as 30% and 25% of cell dry matter when grown on protamylasse or Luria-Bertani medium, respectively. This recombinant strain overcame the bottleneck of the costly arginine provision where it produces about 75% of the CGP obtained from the parent cells grown on mineral medium containing pure arginine as the sole source of carbon. Phosphate starvation is the only known trigger for CGP synthesis in this bacterium, which possesses the PhoB/PhoR phosphate regulon system. Overexpression of phoB caused an 8.6-fold increase in CGP content in comparison to the parent strain at a nonlimiting phosphate concentration.