3-Deoxy-D-manno-octulosonate-8-phosphate synthase catalyzes the C-O bond cleavage of phosphoenolpyruvate.

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.