N-Acetyl-l-glutamate kinase from Escherichia coli: cloning of the gene, purification and crystallization of the recombinant enzyme and preliminary X-ray analysis of the free and ligand-bound forms

The gene for Escherichia coliN-acetyl-l-glutamate kinase (NAGK) was cloned in a plasmid and expressed in E. coli, allowing enzyme purification in three steps. NAGK exhibits high specific activity (1.1 µmol s⁻¹ mg⁻¹), lacks Met1 and forms dimers (shown by cross-linking). Crystals of unliganded NAGK diffract to 2 Å and belong to space group P6₁22 or its enantiomorph P6₅22 (unit-cell parameters a = b = 78.6, c = 278.0 Å) with two monomers in the asymmetric unit. Crystals of NAGK with acetylglutamate and the ATP analogue AMPPNP diffract to 1.8 Å and belong to space group C222₁ (unit-cell parameters a = 60.0, b = 71.9, c = 107.4 Å), with one monomer in the asymmetric unit. NAGK crystallization will allow the determination of proposed structural similarities to carbamate kinase.