OtsAB途径对谷氨酸棒杆菌ATCC 13032胞内海藻糖含量及细胞耐受性的影响

INFLUENCE OF THE CHANGE OF OTSAB PATHWAY ON THE INTRACELLULAR TREHALOSE CONTENT AND CELLULAR TOLERANCE OF CORYNEBACTERIUM GLUTAMICUM ATCC 13032

OtsAB途径是谷氨酸棒杆菌胞内海藻糖3条合成途径中最主要的一条．从野生型谷氨酸棒杆菌（Corynebacterium glutamicum）ATCC13032出发，通过重叠延伸PCR及同源重组技术构建了OtsAB途径缺陷突变株Corynebacterium glutamicum（AotsAB）；利用谷氨酸棒杆菌穿梭表达载体pEC—XK99E构建重组工程菌株Corynebacterium glutamicum（pECotsAB）过量表达同源otsA及otsB基因．考察了野生型、缺陷型及过量表达菌株对于不同渗透压和培养温度的耐受性及该条件下各菌株胞内海藻糖的含量．结果表明：不同培养条件下，Corynebacterium glutamicum（AotsAB）胞内海藻糖含量与同条件野生型相比减少了34．40％～44．96％，对于渗透压和温度的耐受性明显降低；在相同的培养温度（30℃）、不同的渗透压条件下，Corynebacterium glutami．cum（pECotsAB）胞内海藻糖含量与野生型相比增加了14．53％～34．5％；细胞对于温度和渗透压的耐受性随胞内海藻糖含量的增加而提高．

The OtsAB pathway is the most important pathway of the three pathways for intracellular trehalose biosynthesis in Corynebacterium glutamicum. An OtsAB pathway defective mutant Corynebacterium glutamicum AotsAB was constructed from wild - type Corynebacterium glutamicum ATCC 13032 by SOE - PCR and homol- ogous recombination; and OtsA and OtsB genes over- expressing strain were constructed by using Corynebacte- rium glutamicum - E. Colt shuttle expression vector pEC - XK99E at the same time. The intracellular trehalose contents of these two mutants and wild - type strain cultured under different osmotic pressures and at dif- ferent cultivation temperatures were measured, and the tolerance of these strains to osmotic pressure and temperature was researched. The results showed that the defect and over - expression of the OtsAB pathway greatly changed the intracellular trehalose content of Corynebacterium glutamicum, which then might influence the tolerances of cells to osmotic pressure and temperature.