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通过蛋白质工程策略研究Bacillus licheniformisα-amylase的结构和功能快照

Mohsin Shad¹, Muhammad Waheed Akhtar², Muhammad Sajjad²

¹School of Biological Sciences, University of the Punjab, Quaid-e-Azam Campus, Lahore, P.O. 54590, Pakistan; Structural Biology, The Rosalind Franklin Institute, Harwell Science & Innovation Campus, Didcot OX11 0QS, United Kingdom.

International journal of biological macromolecules

|March 19, 2025

概括

巴西菌的alpha-amylase变体被设计为增强粉水解. BLAMCD变种显示活动增加两倍和更高的催化效率,非常适合工业应用.

科学领域:

生物化学生化学
酶学是一种酶学.
蛋白质工程是指蛋白质工程.

背景情况:

阿尔法氨基酶是重要的工业酶,可以分解粉.
巴西菌的α-氨基酶 (BLAM) 是广泛使用的,但可以改进.
了解BLAM变体的结构-功能关系是优化关键.

研究的目的:

为了研究工程化Bacillus licheniformisalpha-amylase (BLAM) 变种的结构和功能性质.
评估N和C端切断和特定突变对BLAM活动和稳定性的影响.
为工业粉加工确定有前途的BLAM变种.

主要方法:

分子对接和MD模拟用于预测残留物稳定性.
用于二次结构和热稳定性分析的循环二元化 (CD) 光谱学.
酶活性测定以确定特定活性和催化效率 (kcat/Km).

主要成果:

工程 BLAM 变种 (BLAMWSP 和 BLAMCD) 在 90°C 显示稳定性.
在90°C时,BLAMCD的特异活性是BLAMWSP (2343.09 ± 0.20 μmol min−1/μmole蛋白) 的两倍 (4237.88 ± 0.66 μmol min−1/μmole蛋白) 的特异活性.
与BLAMWSP (kcat/Km = 51.76 ± 1.76) 相比,BLAMCD的催化效率 (kcat/Km = 114.10 ± 1.41) 和基质亲和力显著提高.

结论:

N和C端的切断和突变提高了BLAM的工业适用性.
由于活性部位的可访问性增加,BLAMCD变异提供了优越的酶性能.
BLAMCD是工业粉液化和化工艺的非常有前途的候选者.

关键词:

细菌类形状is.催化学领域催化学领域蛋白质工程是一种蛋白质工程.α-氨基酶的使用.

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