Yu Xuefei, Li Yanyan, Wang Xiaoyuan
State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, China ; Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, China.
Key Laboratory of Industrial Biotechnology of Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, China.
PLoS One. 2014 Jan 24;9(1):e87550. doi: 10.1371/journal.pone.0087550. eCollection 2014.
In bacteria, threonine dehydratases could convert L-threonine to 2-ketobutyrate. Some threonine dehydratases contain only a catalytic domain, while others contain an N-terminal catalytic domain and a C-terminal regulatory domain composed of one or two ACT-like subdomains. However, the role of the ACT-like subdomain in threonine dehydratases is not clear. Here, nine different bacterial threonine dehydratases were studied. Three of the nine contain no ACT-like subdomain, four of them contain a single ACT-like subdomain, and two of them contain two ACT-like subdomains. The nine genes encoding these threonine dehydratases were individually overexpressed in E. coli BL21(DE3), and the enzymes were purified to homogeneity. Activities of the purified enzymes were analyzed after incubation at different temperatures and different pHs. The results showed that threonine dehydratases with a single ACT-like subdomain are more stable at higher temperatures and a broad range of pH than those without ACT-like subdomain or with two ACT-like subdomains. Furthermore, the specific activity of threonine dehydratases increases with the increase of the number of ACT-like subdomains they contain. The results suggest that the ACT-like subdomain plays an important role in bacterial threonine dehydratases.
在细菌中，苏氨酸脱水酶可将L-苏氨酸转化为2-酮丁酸。一些苏氨酸脱水酶仅包含一个催化结构域，而其他的则包含一个N端催化结构域和一个由一或两个ACT样亚结构域组成的C端调节结构域。然而，ACT样亚结构域在苏氨酸脱水酶中的作用尚不清楚。在此，对九种不同的细菌苏氨酸脱水酶进行了研究。这九种中的三种不含ACT样亚结构域，其中四种含有单个ACT样亚结构域，另外两种含有两个ACT样亚结构域。编码这些苏氨酸脱水酶的九个基因分别在大肠杆菌BL21(DE3)中过表达，并将酶纯化至同质。在不同温度和不同pH下孵育后，分析纯化酶的活性。结果表明，与不含ACT样亚结构域或含有两个ACT样亚结构域的苏氨酸脱水酶相比，含有单个ACT样亚结构域的苏氨酸脱水酶在较高温度和较宽pH范围内更稳定。此外，苏氨酸脱水酶的比活性随着其所含ACT样亚结构域数量的增加而增加。结果表明，ACT样亚结构域在细菌苏氨酸脱水酶中起重要作用。
