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从模块化多基合成酶中获取转移酶域的基底特异性的结构和计算见解

Shuxin Huang¹, Huining Ji¹, Jianting Zheng^1,2

¹State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, China.

The FEBS journal

|June 26, 2024

概括

研究人员在多基合成酶 (PKSs) 中设计了转移酶 (AT) 域,以控制构建块的选择. 这项工作通过了解AT基质特异性来推进新型聚基酸衍生物的工程.

科学领域:

生物化学和分子生物学
自然产品生物合成自然产品生物合成
蛋白质工程是指蛋白质工程.

背景情况:

多基化物是通过多基化合成酶 (PKSs) 合成的天然产物.
在PKS中,乙转移酶 (AT) 域对于在聚化物链组装过程中选择扩展单元至关重要.
工程AT领域提供了一条引入非自然构建块和创建新型多基体结构的途径.

研究的目的:

阐明AT域中从泰洛辛 (TylAT5) 和西诺萨德 (SpnAT8) PKS中选择基质的分子基础.
为了对TylAT5和SpnAT8的基质特异性进行工程,将其转化为马洛尼尔-CoA.
增强对AT域特异性的理解,以改进PKS工程.

主要方法:

确定了TylAT5和SpnAT8AT域的晶体结构.
采用了分子动力学模拟和酶动力学研究.
利用位点定向的突变发生来改变活性位点附近的残留物.

主要成果:

在基质结合口袋内识别了保存的基因 (TylAT5中的TAGH,SpnAT8中的YASH),与它们的原生特异性相关.
局部导向突变发生成功地重新编程了两个AT域,以接受马洛尼尔-CoA作为基质.
在TylAT5的突变增加了对马洛尼尔-CoA的催化活性2.6倍;在SpnAT8的突变减少了基质乱交.

结论:

这项研究提供了关于AT域基质特异性的分子决定因素的详细见解.
重编程AT领域是可行的,为合成生物学的方法在天然产品合成中开辟了道路.
这些发现将促进PKS路径的合理工程,用于生产各种聚基衍生物.

关键词:

乙转移酶是一种转移酶.晶体结构晶体结构分子动力学模拟模拟斯皮诺西尼 (spinosyn) 是一种神经系统.泰洛辛 (tylosin) 是一种药物.

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