热稳定性
氢键
圆二色性
组织谷氨酰胺转胺酶
化学
氨基酸
谷氨酰胺
酶
赖氨酸
立体化学
生物化学
结晶学
有机化学
分子
作者
Xiaoping Song,Hefang Sheng,Yueqiao Zhou,Yin Yu,Yingjiao He,Zihan Wang
摘要
Abstract Microbial transglutaminase (MTG, EC 2.3.2.13) derived from Streptomyces mobaraensis is widely used in the food and pharmaceutical industry because of its ability to synthesize isopeptide bonds between the proteinogenic side chains of glutamine and lysine. The half‐life ( t 1/2 ) of the activated wild‐type enzyme at 60°C is 2 min. To improve the activity and thermostability of MTG for higher temperature application, three variants (Mut1, Mut2, and Mut3) were obtained by combining key amino acid mutations on the basis of previous research results. The best variant Mut2 with a specific combination of five of seven substitutions (S2P–S23V–Y24N–R215A–H289Y) shows a 10‐fold increased half‐life at 60°C ( t 1/2 = 27.6 min), and a 2.4‐fold increased specific enzyme activity (39.3 U/mg). As measured by circular dichroism, the curve of Mut2 was basically the same as that of MTG‐WT. The structural simulation of Mut2 shows that the overall structure is discoid with a crack, but the crack openings are wider than that of MTG‐WT. Furthermore, structural analysis of Mut2 showed that there were seven hydrogen bonds and one π‐anion interaction between Mut2 and its adjacent amino acids, and the number of hydrogen bonds was one more than that of MTG‐WT (six hydrogen bonds).
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