Influence of site-directed mutagenesis of UbiA, overexpression of dxr, menA and ubiE, and supplementation with precursors on menaquinone production in Elizabethkingia meningoseptica
Influence of site-directed mutagenesis of UbiA, overexpression of dxr, menA and ubiE, and supplementation with precursors on menaquinone production in Elizabethkingia meningoseptica
复制标题
UbiA 定点诱变、dxr、menA 和 ubiE 过度表达以及补充前体对 Elizabethkingia meningoseptica 甲基萘醌产生的影响
DOI：
10.1016/j.procbio.2018.01.022
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发表时间：
2018-05-01
影响因子：
4.4
通讯作者：
Yu, Fei
中科院分区：
文献类型：
--
作者：
Liu, Yan;Yang, Zi-ming;Yu, Fei
甲基萘醌（MK）是人体不可缺少的脂溶性维生素，具有广阔的应用前景和潜在的工业生产经济价值。传统上，MK生产已经通过重复突变选择方法开发更好的菌株来改善。然而，由于其随机性和从较大的突变体库中筛选效率低，代谢工程试图改进菌株和发酵过程。据报道，4-羟基苯甲酸异戊烯基转移酶（UbiA）D198定点突变为Ala可使脑膜炎败血性Ellxabethkingla中MK含量增加181%。为了进一步增加，采用了几种系统的代谢工程策略，如共表达限速基因和补充底物前体。共表达dxr和menA并补充丙酮酸钠（SPY）和莽草酸（SA）导致EmUbiA（D198 A）中的MK含量比野生型中没有SPY和SA（CK）高397%。此外，与没有ubiE的这种组合相比，通过dxr、menA和ubiE的共表达以及补充SPY和SA的组合实现了EmUbiA（D198 A）中MK含量的145%增加。总的来说，E.通过削弱泛醌途径，共表达dxr、menA和ubiE，并向培养基中加入SPY和SA，脑膜炎败血症的细胞数增加了11倍以上。
An indispensable fat-soluble vitamin, menaquinone (MK) has promising applications and potential economic value for industrial production. Traditionally, MK production has been improved through the development of better strains by a repeated mutation-selection approach. However, because of its randomness and low screening efficiency from larger mutant libraries, metabolic engineering was attempted to improve strains and the fermentation process. Site-directed mutation of 4-hydroxybenzoate prenyltransferase (UbiA) D198 to Ala has been reported to increase MK content by 181% in Ellxabethkingla meningoseptica. For additional increases, several systematic metabolic engineering strategies, such as co-expression of rate-limiting genes and supplementation with substrate precursors, were employed. Co-expression of dxr and menA and supplementation with sodium pyruvate (SPY) and shikimic add (SA) led to 397% more MK content in EmUbiA (D198A) than that without SPY and SA (CK) in the wild type. Moreover, a 145% increase in the MK content in EmUbiA (D198A) was realized by the combination of co-expression of dxr, menA, and ubiE, and supplementation with SPY and SA compared to that of this combination without ubiE. Overall, MK content in E. meningoseptica increased more than 11 times by weakening the ubiquinone pathway, co-expressing dxr, menA and ubiE, and adding SPY and SA to the medium.