Crystallization and preliminary X-ray analysis of Citrobacter amalonaticus methylaspartate ammonia lyase.
Methylaspartate ammonia lyase (MAL) catalyses the reversible alpha,beta-elimination of ammonia from L-threo-(2S,3S)-3-methylaspartic acid to give mesaconic acid. Crystals of Citrobacter amalonaticus MAL have been obtained by the hanging-drop method of vapour diffusion using ammonium sulfate as the precipitant. Three crystal forms were obtained from identical crystallization conditions, two of which (forms A and B) diffract to high resolution, whilst the third form diffracted poorly. Crystals of form A diffract to beyond 2.1 A and have been characterized as belonging to one of the enantiomorphic space groups P4(1)22 or P4(3)22, with unit-cell parameters a = b = 66.0, c = 233.1 A, alpha = beta = gamma = 90 degrees and a monomer in the asymmetric unit. Crystals of form B diffract to beyond 1.5 A and belong to space group C222, with unit-cell parameters a = 128.3, b = 237.4, c = 65.8 A, alpha = beta = gamma = 90 degrees and a dimer in the asymmetric unit. Determination of the structure of MAL will be an important step in resolving current conflicts concerning the enzyme mechanism which differ between one which places MAL as a member of the superfamily of ammonia lyases whose catalytic activity requires a cofactor formed by post-translational modification of the enzyme and another which links MAL to the enolase superfamily.