EC Number   |
|---|
    2.7.7.9 | - |
| 2.7.7.9 | apo- and UDP-glucose/Mg2+-bound enzyme complexes, hanging drop and sitting drop vapor diffusion methods, protein in 20 mM Tris-HCl pH 7.5 and 0.1 M NaCl, is mixed with 0.1 M sodium acetate trihydrate, pH 4.6, 2 M ammonium sulfate and 0.1 M guanidine-HCl for the apo-enzyme crystals, 22°C, one week. UDP-Glc/Mg2+-bound holo-UGPase is crystallized in 0.1 M HEPES-Na, pH 7.5, 2% PEG 400 and 1.5 M ammonium sulfate containing 10 mM UDP-Glc and 10 mM MgCl2 at 22°C within a month, X-ray diffraction structure determination and analysis at 2.9 A and 2.3 A resolutions, respectively |
| 2.7.7.9 | both in solution and crystal, enzyme forms homooctamers. Association of octamers is mediated by left-handed helices in the C-terminal domains forming a toroidal solenoid structure. The catalytic domains do not directly contact each other, consistent with simple Michaelis-Menten kinetics |
| 2.7.7.9 | detailed comparison between enzyme and thymidylyltransferases |
| 2.7.7.9 | from ammonium sulfate precipitate |
| 2.7.7.9 | hanging-drop vapour-diffusion method at 20°C, crystal structure at resolution of 2.0 A. The crystals reveals the presence of two molecules in the asymmetric unit |
| 2.7.7.9 | in complex with both Mg2+ and UDP-glucose. Residues involved in anchoring the ligand to the active site include the polypeptide chain backbone atoms of Ala20, Gly21, Gly117, Gly180, and Ala214, and the side chains of Glu36, Gln112, Asp143, Glu201, and Lys202. Two magnesium ions are coordinated to the UDP-glucose. An alpha- and a beta-phosphoryl oxygen, three waters, and the side chain of Asp142 ligate the first Mg2+ ion, whereas the second ion is coordinated by an alpha-phosphoryl oxygen and five waters |
| 2.7.7.9 | in complex with glucose 1-phosphate, data from an osmium derivative and a selenomethiomine derivative |
| 2.7.7.9 | in presence of both magnesium and UDP-glucose. Residues anchoring the ligand to the acitve site include polypepetide backbone atoms of A20, G21, G117, G180 and A214 and side chain residues of E36, Q112, D143, E201, and K202. Two magnesium ions coordinate to UDP-glucose |
| 2.7.7.9 | in the D4 octameric structure of the hUGP1-UDP-Glc complex, all subunits have the same overall conformation. The transition of the UGP octamer between the apo- and the product-bound forms is in agreement with the Monod-Wyman-Changeux symmetry model. oligomerzation facilitates an intermolecular stabilization of the sugar moiety in the active site (interlock mechanism), enhances protein stability, enables mild positive cooperativity observed for the octameric wild-type UGP1 towards diphosphate in the reverse reaction, and may allow regulation of the UGP octamer by modification of a single subunit |
