Zhang Sheng, Wilson David B, Ganem Bruce
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
Bioorg Med Chem. 2003 Jul 17;11(14):3109-14. doi: 10.1016/s0968-0896(03)00242-6.
Besides playing a central role in phenylalanine biosynthesis, the bifunctional P-protein in Eschericia coli provides a unique model system for investigating whether allosteric effects can be engineered into protein catalysts using modular regulatory elements. Previous studies have established that the P-protein contains three distinct domains whose functions are preserved, even when separated: chorismate mutase (residues 1-109), prephenate dehydratase (residues 101-285), and an allosteric domain (residues 286-386) for feedback inhibition by phenylalanine. By deleting the prephenate dehydrase domain, a functional chorismate mutase linked directly to the phenylalanine binding domain has been engineered and overexpressed. This manuscript reports the catalytic properties of the mutase in the absence and presence of phenylalanine.
除了在苯丙氨酸生物合成中发挥核心作用外，大肠杆菌中的双功能P蛋白还提供了一个独特的模型系统，用于研究是否可以使用模块化调节元件将变构效应设计到蛋白质催化剂中。先前的研究表明，P蛋白包含三个不同的结构域，即使分开，其功能也能保留：分支酸变位酶（第1-109位氨基酸残基）、预苯酸脱水酶（第101-285位氨基酸残基），以及用于苯丙氨酸反馈抑制的变构结构域（第286-386位氨基酸残基）。通过删除预苯酸脱水酶结构域，构建并过表达了一种直接与苯丙氨酸结合结构域相连的功能性分支酸变位酶。本手稿报道了该变位酶在不存在和存在苯丙氨酸时的催化特性。
