The 5Å resolution crystal structure of adenylosuccinate synthetase from Escherichia coli

Abstract

<p>The technique of X-ray crystallography has been employed to determine the structure of adenylosuccinate synthetase from Escherichia coli to a resolution of 5A. Three crystalline forms of the enzyme have been obtained. The structure determination of the P21 crystal form was pursued due to its excellent diffraction properties and its rapid and reproducible growth. The P21 form crystallizes as a dimer. Unit cell parameters are as follows: a = 73.31(9)A, b = 72.23(7)A, c = 82.87(9)A, [beta] = 108.59°(6). The unit cell volume is 416,700(1,200)A[superscript]3.;A suitable single isomorphous derivative has been obtained with HgI4[superscript]2-. Patterson maps, calculated based on isomorphous and anomalous differences, displayed two peaks corresponding to two heavy atom binding sites per dimer. The heavy atoms are related by a molecular two-fold. The noncrystallographic two-fold axis was located using a self-rotation function and the coordinates of the heavy atom sites.;The intensities were phased based on isomorphous and anomalous differences from the single heavy atom derivative, HgI4[superscript]2-. The electron density map produced from this phase information was not of adequate quality for a chain trace, but it clearly revealed the molecular envelope of the dimer. The dimer displays two-fold symmetry consistent with the results of the self-rotation function and the positions of the heavy atoms. The dimer measures approximately 70A along the b axis and has a maximum radius of 42A in the plane defined by the a and c axes of the unit cell.;Examination of the asymmetric unit reveals density which corresponds to two small domains of the dimer separated by a large, central domain. Moreover, the two crevices created by the interface of the small and large domains serve as sites for the bindings of HgI4[superscript]2-. Mercury has been shown to inactivate the synthetase from E. coli (unpublished results; Dept. of Biochem. and Biophys., Iowa State Univ.). Inhibition may be caused by the binding of mercury to the active site, thus identifying the crevice between the small and large domain as a possibility for the active site.</p>