ANALYSES OF DIFFERENT BINDING MODES OF LIGANDS IN THREE TYPES OF CRYSTALS OF L-ASPARAGINASE FROM E.coli A-1-3KY3598. N. Nandhagopal, M. Hirokawa, N. Tanaka, T. Senda and Y. Mitsui, Department of BioEngineering, Nagaoka University of Technology, Nagaoka, Niigata, 940-21, Japan
Amidohydrolases from Echerichia coli and Erwinia chrysanthemi exhibit a relatively high specificity for asparagine and are referred to as asparaginases. The monoclinic crystal structure of L-asparaginase II from Escherichia coli K12 has been determined at 2.3Å resolution by Swain et al.(1993)1.
The asparaginase from E.coli A-1-3KY35982 was used for crystallization in the present analysis. The molecule is composed of four subunits and the molecular weight is ca. 136 Kda. Three types of crystals, all belonging to a space group P21221, have been obtained, one of them as glutamic acid complexes (Glu-complex) and the other two as aspartic acid complexes (Asp-complex1 and Asp-complex2). For the three types of crystals, the cell parameters were different from each other by up to 15%. In the Glu-complex the loop region in the active site is not clearly resolved in the electron density map. Crystallographically there are two binding sites in each asymmetric unit. In the Asp-complexes, equivalent types of aspartate binding were observed for each of the two binding sites. This may be related to the presence of rigid loop region in each of the two binding sites. Surprisingly, in the Glu-complex, each of the bound glutamate ligands was found to have a different comformation. Detailed analyses of the three types of crystals are in progress.
Comparative studies of the three types of crystal structures and the result of the analyses aiming at clarifying the structural basis of their preferred specificity for asparagine (rather than glutamine) will be presented.
1. Swain, A. et al. (1993) Proc. Natl Acad. Sci. USA, 90, 1974
2. Mitsui, Y. et al. (1978) Acta Crystallogr., A34, 560.
3. Hirokawa, M. et al. (1994) Protein Eng., 7, 1165