Efficient synthesis of 6′-sialyllactose, 6,6′-disialyllactose, and 6′-KDO-lactose by metabolically engineered E. coli expressing a multifunctional sialyltransferase from the Photobacterium sp JT-ISH-224
Efficient synthesis of 6′-sialyllactose, 6,6′-disialyllactose, and 6′-KDO-lactose by metabolically engineered E. coli expressing a multifunctional sialyltransferase from the Photobacterium sp JT-ISH-224
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DOI：
10.1016/j.carres.2010.02.018
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发表时间：
2010-07-02
影响因子：
3.1
通讯作者：
Samain, Eric
中科院分区：
文献类型：
--
作者：
Drouillard, Sophie;Mine, Toshiki;Samain, Eric
我们先前已经报道了通过表达α-（2 -> 3）-唾液酸转移酶的脑膜炎奈瑟氏球菌基因的基因工程大肠杆菌菌株的高细胞密度培养物将乳糖有效转化为3 ′-唾液酸乳糖[Fierfort，N.; Samain，E. J. Biotechnol. 2008，134，261 -265.]。首先尝试使用类似的策略来用表达来自发光杆菌属JT-ISH-224的α-（2 -> 6）-唾液酸转移酶的菌株产生6 '-唾液酸乳糖，导致产生被鉴定为KDO-乳糖（2-酮基-3-脱氧-甘露-辛酰乳糖）的三糖。该结果显示α-（2 -> 6）-唾液酸转移酶能够使用CMP-KDO作为糖供体，并且相对于CMP-Neu 5Ac优先使用CMP-KDO。通过降低唾液酸转移酶基因的表达水平并增加neuABC基因的表达水平，我们已经能够促进6 '-唾液酸乳糖的形成并防止KDO-乳糖的形成。然而，在这种情况下，还产生了第三种乳糖衍生物，其被鉴定为6，6 '-二唾液酸乳糖。主要在乳糖不足的条件下观察到6，6 '-二唾液酸乳糖的形成。另一方面，当用过量的乳糖连续补料培养物时，6 ′-唾液酸乳糖几乎是检测到的唯一产物，并且其最终浓度高于30 g/L培养基。(C)2010爱思唯尔有限公司保留所有权利。
We have previously reported the efficient conversion of lactose into 3'-sialyllactose by high cell density cultures of a genetically engineered Escherichia coli strain expressing the Neisseria meningitidis gene for alpha-(2 -> 3)-sialyltransferase [Fierfort, N.; Samain, E. J. Biotechnol. 2008, 134,261-265.]. First attempts to use a similar strategy to produce 6'-sialyllactose with a strain expressing alpha-(2 -> 6)-sialyltransferase from the Photobacterium sp. JT-ISH-224 led to the production of a trisaccharide that was identified as KDO-lactose (2-keto-3-deoxy-manno-octonyllactose). This result showed that alpha-(2 -> 6)-sialyltransferase was able to use CMP-KDO as sugar donor and preferentially used CMP-KDO over CMP-Neu5Ac. By reducing the expression level of the sialyltransferase gene and increasing that of the neuABC genes, we have been able to favour the formation of 6'-sialyllactose and to prevent the formation of KDO-lactose. However, in this case, a third lactose derivative, which was identified as 6,6'-disialyllactose, was also produced. Formation of 6,6'-disialyllactose was mainly observed under conditions of lactose shortage. On the other hand, when the culture was continuously fed with an excess of lactose, 6'-sialyllactose was almost the only product detected and its final concentration was higher than 30 g/L of culture medium. (C) 2010 Elsevier Ltd. All rights reserved.