Zeng Jing, Guo Jianjun, Tu Yikun, Yuan Lin
1Institute of Microbiology, Jiangxi Academy of Sciences, No. 7777 Changdong Avenue, Nanchang, 330096 Jiangxi Province China.
2School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, 211800 China.
Food Sci Biotechnol. 2019 Aug 26;29(3):409-418. doi: 10.1007/s10068-019-00673-x. eCollection 2020 Mar.
Since the thermoacidophilic raw-starch hydrolyzing -amylase Gt-amy can effectively hydrolyze corn starch under starch liquefaction conditions, it has potential for many industrial applications. To identify the raw starch-binding domain of Gt-amy, a C-terminal domain (CTD)-truncated mutant (Gt-amy-T) was constructed, and its enzymatic properties were compared with Gt-amy. In comparison to CTD of Gt-amy, which could effectively bind corn starch, the Gt-amy-T could not bind to and hydrolyze corn starch under similar conditions. In addition, Gt-amy-T showed significantly lower thermal activity and thermal stability. Using soluble starch as the substrate, the of Gt-amy-T at 80 °C was approximately 77.9% of that of Gt-amy. The half-life of Gt-amy at 80 °C was 3 h, while that of Gt-amy-T was 2 h. These results reveal that the CTD plays a vital role in raw starch binding and degradation by Gt-amy and helps Gt-amy maintain thermal activity and stability.
由于嗜热嗜酸的生淀粉水解α-淀粉酶Gt-amy在淀粉液化条件下能有效水解玉米淀粉，因此它在许多工业应用中具有潜力。为了鉴定Gt-amy的生淀粉结合结构域，构建了一个C末端结构域（CTD）截短的突变体（Gt-amy-T），并将其酶学性质与Gt-amy进行比较。与能有效结合玉米淀粉的Gt-amy的CTD相比，Gt-amy-T在相似条件下不能结合和水解玉米淀粉。此外，Gt-amy-T的热活性和热稳定性显著降低。以可溶性淀粉为底物，Gt-amy-T在80℃时的活性约为Gt-amy的77.9%。Gt-amy在80℃的半衰期为3小时，而Gt-amy-T的半衰期为2小时。这些结果表明，CTD在Gt-amy结合和降解生淀粉过程中起着至关重要的作用，并有助于Gt-amy保持热活性和稳定性。
