Structure-based approach for binding site identification on AmpC β-lactamase
Structure-based approach for binding site identification on AmpC β-lactamase
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DOI：
10.1021/jm020002p
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发表时间：
2002-07-18
影响因子：
7.3
通讯作者：
Shoichet, BK
中科院分区：
文献类型：
--
作者：
Powers, RA;Shoichet, BK
β-内酰胺酶是对β-内酰胺类抗生素最普遍的耐药机制，对公众健康构成越来越大的威胁。已经确定了几种β-内酰胺酶的结构，使这种酶成为基于结构的药物设计的一个有吸引力的目标。为了方便C类β-内酰胺酶AmpC的抑制剂设计，通过实验和计算的方法确定了该酶上的结合热点。实验测定了四种硼酸抑制剂与较高分辨率(1.72埃)天然载脂蛋白的配合物中AMPC的X-射线晶体结构。除了先前确定的AmpC在与其他五种硼酸抑制剂和四种β-内酰胺类药物的络合物中的结构外，还鉴定了共同的结合部位。在计算上，使用GRID、MCSS和X-SITE程序预测AmpC上潜在的结合位点热点。从晶体结构中确定了几个一致的结合部位。通过β-内酰胺的R1侧链上的羰基氧与保守的Asn152的Ndelta2原子之间的相互作用，确定了一个酰胺识别位点。令人惊讶的是，这个位置还识别芳基硼酸的芳环，似乎与Asn152形成四极-偶极相互作用。高度保守的“氧阴离子”空穴定义了一个既能识别羰基又能识别羟基的位置。通过硼酸中的O2羟基确定了一个羟基结合部位，它与Tyr150和保守的水形成氢键。Leu119和Leu293形成疏水中心。通过与Asn346和Arg349相互作用的β-内酰胺类化合物中普遍存在的C3(4)羧酸盐，确定了一个羧酸结合部位。在大多数结构中观察到的有序水识别出四个水位；这些水与AMPC形成广泛的氢键网络，偶尔还与配体形成氢键网络。计算程序的预测与实验观察到的结合位点有一定的相关性。有几个结合位点没有被预测，但提出了新的结合位点。综上所述，构建了在AMPC上发现的结合位点热点的地图，以及关于每个位点识别的功能的信息。这一图谱可能有助于针对AmpC进行基于结构的抑制剂设计。
beta-Lactamases are the most widespread resistance mechanism to beta-lactam antibiotics and are an increasing menace to public health. Several beta-lactamase structures have been determined, making this enzyme an attractive target for structure-based drug design. To facilitate inhibitor design for the class C beta-lactamase AmpC, binding site "hot spots" on the enzyme were identified using experimental and computational approaches. Experimentally, X-ray crystal structures of AmpC in complexes with four boronic acid inhibitors and a higher resolution (1.72 Angstrom) native apo structure were determined. Along with previously determined structures of AmpC in complexes with five other boronic acid inhibitors and four beta-lactams, consensus binding sites were identified. Computationally, the programs GRID, MCSS, and X-SITE were used to predict potential binding site hot spots on AmpC. Several consensus binding sites were identified from the crystal structures. An amide recognition site was identified by the interaction between the carbonyl oxygen in the R1 side chain of beta-lactams and the atom Ndelta2 of the conserved Asn152. Surprisingly, this site also recognizes the aryl rings of arylboronic acids, appearing to form quadrupole-dipole interactions with Asn152. The highly conserved "oxyanion" hole defines a site that recognizes both carbonyl and hydroxyl groups. A hydroxyl binding site was identified by the O2 hydroxyl in the boronic acids, which hydrogen bonds with Tyr150 and a conserved water. A hydrophobic site is formed by Leu119 and Leu293. A carboxylate binding site was identified by the ubiquitous C3(4) carboxylate of the beta-lactams, which interacts with Asn346 and Arg349. Four water sites were identified by ordered waters observed in most of the structures; these waters form extensive hydrogen-bonding networks with AmpC and occasionally the ligand. Predictions by the computational programs showed some correlation with the experimentally observed binding sites. Several sites were not predicted, but novel binding sites were suggested. Taken together, a map of binding site hot spots found on AmpC, along with information on the functionality recognized at each site, was constructed. This map may be useful for structure-based inhibitor design against AmpC.