2E25
The Crystal Structure of the T109S mutant of E. coli Dihydroorotase complexed with an inhibitor 5-fluoroorotate
2E25 の概要
| エントリーDOI | 10.2210/pdb2e25/pdb |
| 関連するPDBエントリー | 1XGE |
| 分子名称 | Dihydroorotase, ZINC ION, 5-FLUORO-2,6-DIOXO-1,2,3,6-TETRAHYDROPYRIMIDINE-4-CARBOXYLIC ACID, ... (4 entities in total) |
| 機能のキーワード | tim barrel, hydrolase |
| 由来する生物種 | Escherichia coli |
| タンパク質・核酸の鎖数 | 1 |
| 化学式量合計 | 39061.96 |
| 構造登録者 | |
| 主引用文献 | Lee, M.,Maher, M.J.,Guss, J.M. Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-fluoroorotate: catalytic activity is reflected by the crystal form Acta Crystallogr.,Sect.F, 63:154-161, 2007 Cited by PubMed Abstract: Crystals of a single-point mutant (T109S) of Escherichia coli dihydroorotase (DHOase) with diminished activity grown in the presence of L-dihydroorotate (L-DHO) are tetragonal, with a monomer in the asymmetric unit. These crystals are extremely unstable and disintegrate shortly after formation, which is followed by the growth of orthorhombic crystals from the remnants of the tetragonal crystals or at new nucleation sites. Orthorhombic crystals, for which a structure has previously been reported [Thoden et al. (2001), Biochemistry, 40, 6989-6997; Lee et al. (2005), J. Mol. Biol. 348, 523-533], contain a dimer of DHOase in the asymmetric unit; the active site of one monomer contains the substrate N-carbamyl-L-aspartate (L-CA-asp) and the active site of the other monomer contains the product of the reaction, L-DHO. In the subunit with L-DHO in the active site, a surface loop (residues 105-115) is 'open'. In the other subunit, with L-CA-asp in the active site, the loop folds inwards, forming specific hydrogen bonds from the loop to the L-CA-asp. The tetragonal crystal form can be stabilized by crystallization in the presence of the inhibitor 5-fluoroorotate (FOA), a product (L-DHO) mimic. Crystals of the complex of T109S DHOase with FOA are tetragonal, space group P4(1)2(1)2, with unit-cell parameters a = b = 72.6, c = 176.1 A. The structure has been refined to R and R(free) values of 0.218 and 0.257, despite severe anisotropy of the diffraction. In this structure, the flexible loops are both in the 'open' conformation, which is consistent with FOA, like L-DHO, binding at both sites. The behaviour of the T109S mutant crystals of DHOase in the presence of L-DHO is explained by initial binding of L-DHO to both subunits, followed by slow conversion to L-CA-asp, with consequent movement of the flexible loop and dissolution of the crystals. Orthorhombic crystals are then able to grow in the presence of L-DHO and L-CA-asp. PubMed: 17329804DOI: 10.1107/S1744309107004009 主引用文献が同じPDBエントリー |
| 実験手法 | X-RAY DIFFRACTION (2.7 Å) |
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