Gross quaternary changes in aspartate carbamoyltransferase are induced by the binding of N-(phosphonacetyl)-L-aspartate: A 3.5-A resolution study.

The three-dimensional structure of the complex of N-(phosphonacetyl)-L-aspartate with aspartate carbamoyltransferase (carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2) has been determined to a nominal resolution of 3.5 A by single-crystal x-ray diffraction methods. Initial phases were obtained by the method of "molecular tectonics": beginning with the structure of the CTP-protein complex, the domains of the catalytic and regulatory chains were manipulated as separate rigid bodies. The resulting coordinates were used to calculate an electron density map, which was then back transformed to give a set of calculated amplitudes and phases. Using all observed data, we obtained a crystallographic R factor between observed and calculated amplitudes Fo and Fc of 0.46. An envelope was then applied to a 2Fo - Fc map and the density was averaged across the molecular twofold axis. Two cycles of averaging yielded an R factor of 0.25. In this complex, we find that the two catalytic trimers have separated from each other along the threefold axis by 11-12 A and have rotated in opposing directions around the threefold axis such that the total relative reorientation is 8-9 degrees. This rotation places the trimers in a more nearly eclipsed configuration. In addition, two domains in a single catalytic chain have changed slightly their spatial relationship to each other. Finally, the two chains of one regulatory dimer have rotated 14-15 degrees around the twofold axis, and the Zn domains have separated from each other by 4 A along the threefold axis. These movements enlarge the central cavity of the molecule and allow increased accessibility to this cavity through the six channels from the exterior surface of the enzyme.