Screening, expression, purification and characterization of CoA-transferases for lactoyl-CoA generation
Screening, expression, purification and characterization of CoA-transferases for lactoyl-CoA generation
复制标题
用于乳酰辅酶 A 生成的辅酶 A 转移酶的筛选、表达、纯化和表征
DOI：
10.1007/s10295-019-02174-6
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发表时间：
2019-07-01
影响因子：
3.4
通讯作者：
Chen, Tao
中科院分区：
文献类型：
--
作者：
Zhang, Xiaoxia;Mao, Yufeng;Chen, Tao
乳酰辅酶A是生物可降解和生物相容性乳酸酯共聚物生物合成的关键，具有广泛的应用。然而，关于乙酰辅酶A：乳酸辅酶A转移酶（ALCT）的报道很少。为了开发新的ALCT，基于丙酸梭菌和埃氏巨球菌的CoA-转移酶进行氨基酸序列相似性搜索。两个已知的和三个新的酶的表达，纯化和表征。鉴定了三种新的ALCT，分别来自Megaspaera sp. DISK 18、乳酸发酵梭菌An 75和厚壁菌属细菌CAG：466。来自埃氏巨球菌的ME-PCT对乙酰辅酶A（264.22s（-1）mM（-1））和d-乳酸（84.18s（-1）mM（-1））都具有最高的催化效率，具有宽的活性温度范围和良好的稳定性。因此，这项研究提供了新的和有效的酶乳酰辅酶A的生产。据我们所知，这是第一份关于ALCT系统挖掘的报告，它为依赖这些酶的途径的工程设计提供了有价值的新工具。
Lactoyl-CoA is critical for the biosynthesis of biodegradable and biocompatible lactate-based copolymers, which have wide applications. However, reports on acetyl-CoA: lactate CoA-transferases (ALCTs) are rare. To exploit novel ALCTs, amino acid sequence similarity searches based on the CoA-transferases from Clostridium propionicum and Megasphaera elsdenii were conducted. Two known and three novel enzymes were expressed, purified and characterized. Three novel ALCTs were identified, one each from Megasphaera sp. DISK 18, Clostridium lactatifermentans An75 and Firmicutes bacterium CAG: 466. ME-PCT from Megasphaera elsdenii had the highest catalytic efficiency for both acetyl-CoA (264.22s(-1)mM(-1)) and d-lactate (84.18s(-1)mM(-1)) with a broad temperature range for activity and good stability. This study, therefore, offers novel and efficient enzymes for lactoyl-CoA generation. To our best knowledge, this is the first report on the systematic mining of ALCTs, which offers valuable new tools for the engineering of pathways that rely on these enzymes.