Kinetic and Structural Studies of E. Coli Dihydrodipicolinate Synthase and Meso-diaminopimelate

Abstract

Lysine is a member of the aspartate family of amino acids. In general, there are two different pathways for the biosynthesis of lysine; the diaminopimelate (DAP) pathway is found in all bacteria, algae, and plants, and the ?-aminoadipic acid pathway seen in fungi and euglenoids. Neither pathway is found in mammals. Dihydrodipicolinate synthase catalyzes the first step in the DAP pathway for the biosynthesis of L-lysine. The enzyme is feedback inhibited allosterically by L-lysine which reduces enzyme activity by ninety percent compared to the uninhibited activity. The kinetic mechanism for DHDPS is ping pong with pyruvate binding first to apo-enzyme followed by generation of a Schiff’s base between pyruvate and K161. Subsequent loss of a proton from the ?-methyl group of the bound pyruvate leads to formation of an enamine intermediate. Based on protein docking studies several substrate and transition analogues of DHDPS have been designed. Kinetic and structural studies was performed on meso-diaminopimelate (meso-DAP). Meso-DAP is a weak activator of DHDPS, increasing the rate of the reaction by 20%. Crystals of dihydrodipicolinate synthase co-complexed with meso-DAP formed in PEG 3350, sodium tartrate, and HEPES at pH 7.5. The diffraction data will be collected at the OU X-ray facility. Structural studies elucidate the binding site for meso-DAP. *This research is funded by Institutional Development Award (IDeA) from the National Institutes of Health (P20GM1034