在人类肠道中的普卢兰酶中N-终端域的功能性作用 Lactobacillus acidophilus
Yu Wang1, Birte Svensson1, Bernard Henrissat2
1Enzyme and Protein Chemistry, Department of Biotechnology and Biomedicine, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark.
Journal of agricultural and food chemistry
|December 6, 2023
概括
在Lactobacillus acidophilus pullulanase中的N端域对于粉结合和酶稳定性至关重要. 删除这些域,特别是CBM41和DUF,会显著影响基板识别和热稳定性.
科学领域:
- 酶学 是一种酶学.
- 生物化学 生物化学
- 微生物学 微生物学
背景情况:
- 普卢兰酶是具有N端域 (NTD) 的α-葡萄糖分支酶,包括碳水化合物结合模块 (CBM) 和未知功能的域 (DUF).
- 了解NTD在酶活性和稳定性中的作用对于酶工程和应用至关重要.
研究的目的:
- 调查CBM41和DUF在Lactobacillus acidophilus NCFM pullulanase (LaPul) 中的特定作用.
- 确定NTD如何影响LaPul的基质结合,催化效率和热稳定性.
主要方法:
- 两种截断的LaPul变种的重组生产: Δ41-LaPul (缺乏CBM41) 和 Δ(41+DUF) -LaPul (缺乏CBM41和DUF).
- 界面动力学测试用于测量粉颗粒上的酶攻击部位.
- 酶动力学 (kcat,KM) 和热稳定性 (化温度) 的测量.
主要成果:
- 拉普尔在粉颗粒上识别的酶攻击部位明显多于截断的变种.
- CBM41主要负责拉普尔对粉颗粒和β-cyclopodextrin的高度亲和力.
- DUF对于LaPul的热稳定性至关重要,因为Δ{\\displaystyle \\Delta }41+DUFs) -LaPul的化温度低12°C.
- 与LaPul相比,Δ41-LaPul的催化效率发生了变化 (kcat更高,KM更高).
结论:
- 在拉普尔的NTD中,CBM41和DUF在基质识别和酶稳定中发挥着独特而至关重要的作用.
- NTDs的切断影响了基质结合亲和力和催化性能.
- 这些发现提供了关于pullulanases的结构-功能关系和酶工程潜力的见解.
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