Hedstrom L, Abeles R
Graduate Department of Biochemistry, Brandeis University, Waltham, MA 02254.
Biochem Biophys Res Commun. 1988 Dec 15;157(2):816-20. doi: 10.1016/s0006-291x(88)80322-x.
The mechanism of 3-deoxy-D-manno-octulosonate-8-phosphate (KDO8P) synthase was investigated. When [18O]-PEP specifically labeled in the enolic oxygen is a substrate for KDO8P synthase, the 18O is recovered in Pi. This indicates that the KDO8P synthase reaction proceeds with C-O bond cleavage of PEP similar to that observed in the 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase catalyzed condensation of PEP and erythrose-4-phosphate (1). No evidence for a covalent enzyme-PEP intermediate could be obtained. No [32P]-Pi exchange into PEP nor scrambling of bridge 18O to non-bridging positions in [18O]-PEP was observed in the presence or absence of arabinose-5-phosphate or its analog ribose-5-phosphate. Bromopyruvate inactivated KDO8P synthase in a time dependent process. It is likely that bromopyruvate reacts with a functional group at the PEP binding site since PEP, but not arabinose-5-phosphate, protects against inactivation.
对3-脱氧-D-甘露辛酮糖酸-8-磷酸（KDO8P）合酶的作用机制进行了研究。当在烯醇式氧中特异性标记的[18O]-PEP作为KDO8P合酶的底物时，18O在Pi中被回收。这表明KDO8P合酶反应是通过PEP的C-O键断裂进行的，类似于在3-脱氧-D-阿拉伯庚酮糖酸-7-磷酸合酶催化的PEP与赤藓糖-4-磷酸的缩合反应中观察到的情况（1）。未获得共价酶-PEP中间体的证据。在存在或不存在阿拉伯糖-5-磷酸或其类似物核糖-5-磷酸的情况下，均未观察到[32P]-Pi与PEP的交换，也未观察到[18O]-PEP中桥连18O向非桥连位置的重排。溴丙酮酸在一个时间依赖性过程中使KDO8P合酶失活。溴丙酮酸可能与PEP结合位点处的一个官能团发生反应，因为PEP而非阿拉伯糖-5-磷酸可防止失活。
