The thermodynamic signature of ligand binding to histone deacetylase-like amidohydrolases is most sensitive to the flexibility in the L2-loop lining the active site pocket.
The thermodynamic signature of ligand binding to histone deacetylase-like amidohydrolases is most sensitive to the flexibility in the L2-loop lining the active site pocket.
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DOI：
10.1016/j.bbagen.2017.04.001
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发表时间：
2017-07
期刊：
影响因子：
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通讯作者：
C. Meyners;Andreas Krämer;Özkan Yildiz;F. Meyer‐Almes
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文献类型：
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作者：
C. Meyners;Andreas Krämer;Özkan Yildiz;F. Meyer‐Almes
背景配体-蛋白质结合的热力学驱动力的分析被认为是选择和优化活性化合物成为候选药物的关键组成部分。从等温滴定量热法 (ITC) 推导出来的结合焓通常被解释为假设配体与靶蛋白的一种构象的单步结合。尽管在许多情况下是成功的，但这些假设在许多情况下都是对现实的过度简化的近似，具有灵活的蛋白质和复杂的结合机制。蛋白质柔性与配体结合的热力学特征之间的关系尚未得到充分研究。方法结合定向诱变、X 射线晶体学、酶动力学和 ITC 方法来剖析环柔性对热力学的影响以及配体与组蛋白脱乙酰酶 (HDAC) 样酰胺水解酶结合的机制。结果一般的配体-蛋白质结合机制包括一个需要能量的大门打开步骤，然后是物理结合。 HDAC 类酰胺水解酶中 L2 环的灵活性增加，有利于配体进入结合袋，导致主要由焓驱动的复合物形成。结论该研究提供了证据，证明了活性位点通道附近的灵活性对该机制的重要性，并观察到 HDAC 类酶中分子识别的热力学驱动力。一般意义在设计更好的候选药物时，应更多地关注与结合袋相邻区域的灵活性或延展性。所提出的案例研究还表明，应谨慎解释观察到的蛋白质-配体系统的结合焓，因为更复杂的结合机制可能会掩盖潜在药物相似性的重要性。
BackgroundThe analysis of the thermodynamic driving forces of ligand-protein binding has been suggested to be a key component for the selection and optimization of active compounds into drug candidates. The binding enthalpy as deduced from isothermal titration calorimetry (ITC) is usually interpreted assuming single-step binding of a ligand to one conformation of the target protein. Although successful in many cases, these assumptions are oversimplified approximations of the reality with flexible proteins and complicated binding mechanism in many if not most cases. The relationship between protein flexibility and thermodynamic signature of ligand binding is largely understudied.MethodsDirected mutagenesis, X-ray crystallography, enzyme kinetics and ITC methods were combined to dissect the influence of loop flexibility on the thermodynamics and mechanism of ligand binding to histone deacetylase (HDAC)-like amidohydrolases.ResultsThe general ligand-protein binding mechanism comprises an energetically demanding gate opening step followed by physical binding. Increased flexibility of the L2-loop in HDAC-like amidohydrolases facilitates access of ligands to the binding pocket resulting in predominantly enthalpy-driven complex formation.ConclusionsThe study provides evidence for the great importance of flexibility adjacent to the active site channel for the mechanism and observed thermodynamic driving forces of molecular recognition in HDAC like enzymes.General significanceThe flexibility or malleability in regions adjacent to binding pockets should be given more attention when designing better drug candidates. The presented case study also suggests that the observed binding enthalpy of protein-ligand systems should be interpreted with caution, since more complicated binding mechanisms may obscure the significance regarding potential drug likeness.