Predicting and harnessing protein flexibility in the design of species-specific inhibitors of thymidylate synthase
Predicting and harnessing protein flexibility in the design of species-specific inhibitors of thymidylate synthase
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DOI：
10.1016/s1074-5521(01)00067-9
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发表时间：
2001-10-01
影响因子：
--
通讯作者：
Stroud, RM
中科院分区：
文献类型：
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作者：
Fritz, TA;Tondi, D;Stroud, RM
背景：蛋白质对配体结合的可塑性消除了刚性受体位点的概念。因此，计算对接本身错过了重要的前瞻性药物设计线索。使用基于计算机的筛选、随后并行合成精制和酶测定的组合开发了必需酶胸苷酸合酶（TS）的细菌特异性抑制剂[Tondi等（1999）Chem.Biol.6，319-331]。特异性是通过蛋白质可塑性实现的，尽管物种之间的酶具有非常高的序列保守性。结果如下：合成的最有效的抑制剂，N，O-二丹酰-L-酪氨酸（DDT），结合干酪乳杆菌TS（LcTS）的亲和力高35倍，大肠杆菌TS（EcTS）的亲和力高24倍，比人TS（hTS）。为了揭示这种特异性的分子基础，我们确定了与DDT和2 '-脱氧尿苷-5'-单磷酸（dUMP）复合的EcTS的晶体结构。2.0埃的结构表明，DDT结合EcTS的构象没有预测的分子对接研究和实质上不同于其他TS抑制剂。DDT的结合是伴随着大的重排的蛋白质附近和远端的酶的活性位点与运动的Ca碳高达6 A相对于其他三元复合物。这种蛋白质的可塑性导致与DDT的新的相互作用，包括形成氢键和货车德瓦尔斯相互作用，以在细菌TS中保守的残基，但不是hTS，并假设占DDT的特异性。构象滴滴涕采取结合EcTS时，解释了其他几个LcTS抑制剂的活性，平行合成的DDT，这表明DDT结合到这两种酶在类似的方向。结论：戏剧性的蛋白质重排涉及的主要和侧链原子发挥了重要作用，在识别DDT的EcTS和突出的重要性，将蛋白质的可塑性在药物设计。EcTS/dUMP/DDT复合物的晶体结构是开发针对致病细菌物种的更有选择性的TS抑制剂的模型系统。晶体结构还提出了一个通用公式，用于识别TS和其他酶的区域，这些区域可以被视为灵活的，以帮助药物发现的计算方法。(C)2001爱思唯尔科技有限公司。保留所有权利。
Background: Protein plasticity in response to ligand binding abrogates the notion of a rigid receptor site. Thus, computational docking alone misses important prospective drug design leads. Bacterial-specific inhibitors of an essential enzyme, thymidylate synthase (TS), were developed using a combination of computer-based screening followed by in-parallel synthetic elaboration and enzyme assay [Tondi et al. (1999) Chem. Biol. 6, 319-331]. Specificity was achieved through protein plasticity and despite the very high sequence conservation of the enzyme between species. Results: The most potent of the inhibitors synthesized, N,O-didansyl-L-tyrosine (DDT), binds to Lactobacillus casei TS (LcTS) with 35-fold higher affinity and to Escherichia coli TS (EcTS) with 24-fold higher affinity than to human TS (hTS). To reveal the molecular basis for this specificity, we have determined the crystal structure of EcTS complexed with DDT and 2'-deoxyuridine-5'-monophosphate (dUMP). The 2.0 Angstrom structure shows that DDT binds to EcTS in a conformation not predicted by molecular docking studies and substantially differently than other TS inhibitors. Binding of DDT is accompanied by large rearrangements of the protein both near and distal to the enzyme's active site with movement of Ca carbons up to 6 A relative to other ternary complexes. This protein plasticity results in novel interactions with DDT including the formation of hydrogen bonds and van der Waals interactions to residues conserved in bacterial TS but not hTS and which are hypothesized to account for DDT's specificity. The conformation DDT adopts when bound to EcTS explains the activity of several other LcTS inhibitors synthesized in-parallel with DDT suggesting that DDT binds to the two enzymes in similar orientations. Conclusions: Dramatic protein rearrangements involving both main and side chain atoms play an important role in the recognition of DDT by EcTS and highlight the importance of incorporating protein plasticity in drug design. The crystal structure of the EcTS/dUMP/DDT complex is a model system to develop more selective TS inhibitors aimed at pathogenic bacterial species. The crystal structure also suggests a general formula for identifying regions of TS and other enzymes that may be treated as flexible to aid in computational methods of drug discovery. (C) 2001 Elsevier Science B.V. All rights reserved.