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 coli N-acetyl-L-glutamate kinase (NAGK) was cloned in a plasmid and expressed in E. coli, allowing enzyme purification in thr ee steps. NAGK exhibits high specific activity (1.1 mu mol s(-1) mg(-1)), l acks Met1 and forms dimers (shown by crosslinking). Crystals of unliganded NAGK diffract to 2 Angstrom and belong to space group P6(1)22 or its enanti omorph P6(5)22 (unit-cell parameters a = b = 78.6, c = 278.0 Angstrom) with two monomers in the asymmetric unit. Crystals of NAGK with acetylglutamate and the ATP analogue AMPPNP diffract to 1.8 Angstrom and belong to space g roup C222(1) (unit-cell parameters a = 60.0, b = 71.9, c = 107.4 Angstrom), with one monomer in the asymmetric unit. NAGK crystallization will allow t he determination of proposed structural similarities to carbamate kinase.