Spinach leaf dihydrodipicolinate synthase: Partial purification and characterization

The first enzyme unique to lysine biosynthesis in higher plants, dihydrodipicolinate synthase, has been partially purified from spinach leaves, using ion exchange chromatography, hydrophobic interaction chromatography and gel filtration. The spinach enzyme is moderately stable to short-term exposure to heat, in contrast to the pea leaf enzyme, but is unstable on storage even at ?20°. Thiol reagents interfere with the calorimetric assay used, and so cannot be routinely used to stabilize the enzyme, which has an active sulphydryl group. The MW of the enzyme is 115000 (gel filtration). Lysine is a potent inhibitor with an I_(0.5) of 2OμM, whilst the lysine analogue S-β-aminoethylcysteinc has an I_(0.5) of 400 μM. The Kt´_m for aspartic-β-semialdehyde was determined to be 1.4mM, but this compound demonstrated marked substrate inhibition at concentrations above 7 mM, increasing the apparent S_(0.5)for the second substrate, pyruvate.