Parker E J, Bulloch E M, Jameson G B, Abell C
Centre for Structural Biology, Institute of Fundamental Sciences, Massey University, Palmerston North, New Zealand.
Biochemistry. 2001 Dec 11;40(49):14821-8. doi: 10.1021/bi010928j.
3-Deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAH7PS, EC 4.1.2.15) catalyzes the condensation of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to give DAH7P via an ordered sequential mechanism. In the absence of PEP (the first substrate to bind), E4P binds covalently to the phenylalanine-sensitive DAH7PS of Escherichia coli, DAH7PS(Phe), deactivating the enzyme. Activity is restored on addition of excess PEP but not if deactivation was carried out in the presence of sodium cyanoborohydride. Electrospray mass spectrometry indicates that a single E4P is bound to the protein. These data are consistent with a slow, reversible Schiff base reaction of the aldehydic functionality of E4P with a buried lysine. Molecular modeling indicates that Lys186, a residue at the base of the substrate-binding cavity involved in hydrogen bonding with PEP, is well placed to react with E4P forming an imine linkage that is substantially protected from solvent water.
3-脱氧-D-阿拉伯庚酮糖酸-7-磷酸合酶（DAH7PS，EC 4.1.2.15）通过有序顺序机制催化磷酸烯醇丙酮酸（PEP）与4-磷酸赤藓糖（E4P）缩合生成DAH7P。在没有PEP（第一个结合的底物）的情况下，E4P与大肠杆菌的苯丙氨酸敏感型DAH7PS即DAH7PS（Phe）共价结合，使酶失活。加入过量PEP后活性恢复，但如果在氰基硼氢化钠存在下进行失活则不会恢复。电喷雾质谱表明单个E4P与蛋白质结合。这些数据与E4P的醛基官能团与一个埋藏的赖氨酸发生缓慢、可逆的席夫碱反应一致。分子模拟表明，Lys186是底物结合腔底部参与与PEP形成氢键的一个残基，它与E4P反应形成亚胺键的位置良好，该亚胺键基本免受溶剂水的影响。
