D-alanine:D-alanine ligase: Phosphonate and phosphinate intermediates with type and the Y216F mutant
D-alanine:D-alanine ligase: Phosphonate and phosphinate intermediates with type and the Y216F mutant
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DOI：
10.1021/bi962431t
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发表时间：
1997-03-04
期刊：
影响因子：
2.9
通讯作者：
Knox, JR
中科院分区：
文献类型：
--
作者：
Fan, C;Park, IS;Knox, JR
测定了D-丙氨酸：大肠杆菌ddlB基因的D-丙氨酸连接酶与D-丙氨酸-D-α-羟基丁酸酯络合物的晶体结构和Y216F突变连接酶与D-丙氨酸-D-丙氨酸络合物的结构，其分辨率分别为2.2Angstrom和1.9Angstrom，R因子分别为0.156和0.158。在每个复合体中，抑制剂与三磷酸腺苷反应生成ADP和紧密结合的磷酸化过渡态中间体。将这两种结构与已知的野生型连接酶亚磷酸中间体的晶体结构进行比较，发现没有重大的构象变化，但B因子表明覆盖结合位点的环的迁移率存在差异。两种抑制剂对Y216F突变体的抑制作用较弱，部分原因是环间氢键的丧失。类似的机制可能解释了对VANA抑制较差的原因，VANA是由万古霉素耐药肠球菌产生的同源D-丙氨酸：D-乳酸连接酶。
The crystallographic structure of the D-alanine:D-alanine ligase of the ddlB gene of Escherichia coli complexed with a D-Ala-D-a-hydroxybutyrate phosphonate and the structure of the Y216F mutant ligase complexed with a D-Ala-D-Ala phosphinate have been determined to 2.2 and 1.9 Angstrom resolution, respectively, and refined to R factors of 0.156 and 0.158. In each complex the inhibitor has reacted with ATP to produce ADP and a tight-binding phosphorylated transition state intermediate. Comparison of these two structures with the known crystal structure of the phosphinate intermediate of the wild-type ligase shows no major conformational changes, but B factors indicate differences in mobility of loops covering the binding site. The weaker inhibition of the Y216F mutant by both inhibitors is thought to be due in part to the loss of an interloop hydrogen bond. A similar mechanism may account for poor inhibition of VanA, the homologous D-Ala:D-lactate ligase produced by vancomycin-resistant enterococci.