Departament of Chemistry, Center of Exact Sciences, State University of Londrina, Londrina, PR, Brazil.
Departament of Chemistry, Center of Exact Sciences, State University of Londrina, Londrina, PR, Brazil.
Chem Biol Interact. 2022 Sep 25;365:110045. doi: 10.1016/j.cbi.2022.110045. Epub 2022 Jul 16.
We report the investigation of hydantoins and thiohydantoins derived from L and d-amino acids as inhibitors against the Canavalia ensiformis urease (CEU). The biochemical in vitro assay against CEU revealed a promising inhibitory potential for most thiohydantoins with six of them showing %I higher than the reference inhibitor thiourea (56.5%). In addition, thiohydantoin derived from l-valine, 1b, as well as the hydantoin 2d, derived from l-methionine, were identified as the most potent inhibitors with %I = 90.5 and 85.9 respectively. Enzyme kinetic studies demonstrated a mixed and uncompetitive inhibition profile for these compounds with K values of 0.42 mM for 1b and 0.99 mM for 2d. These kinetic parameters, obtained from traditional colorimetric assay, were strictly related to the K values measured spectroscopically by the Saturation Transfer Difference (STD) technique for the urease complex. STD was also used to evince the moieties of the ligands responsible for the binding with the enzyme. Molecular docking studies showed that the thiohydantoin and hydantoin rings can act as a pharmacophoric group due to their binding affinity by hydrogen bonding interactions with critical amino acid residues in the enzyme active and/or allosteric site. These findings agreed with the experimental alpha values, demonstrating that 1b has affinity by free enzyme, and 2d derivative, an uncompetitive inhibitor, has great binding affinity at the allosteric site. The results for the thiohydantoin 1a, derived from d-valine, demonstrated a drastic stereochemical influence on inhibition, kinetics, and binding parameters in comparison to its enantiomer 1b.
我们报告了对来自 L 和 d-氨基酸的海因衍生物和硫代海因衍生物作为刀豆脲酶（CEU）抑制剂的研究。对 CEU 的生化体外测定显示，大多数硫代海因具有有希望的抑制潜力，其中有 6 种的%I 高于参考抑制剂硫脲（56.5%）。此外，衍生自 l-缬氨酸的硫代海因 1b 以及衍生自 l-蛋氨酸的海因 2d 被鉴定为最有效的抑制剂，%I 分别为 90.5 和 85.9。酶动力学研究表明，这些化合物具有混合和非竞争性抑制特征，对于这些化合物，K 值分别为 1b 的 0.42 mM 和 2d 的 0.99 mM。从传统比色测定获得的这些动力学参数与通过饱和转移差异（STD）技术在酶复合物中光谱测量的 K 值严格相关。STD 还用于证明配体的部分负责与酶结合。分子对接研究表明，硫代海因和海因环可以作为药效团，因为它们通过与酶的活性和/或变构部位的关键氨基酸残基的氢键相互作用而具有结合亲和力。这些发现与实验的α值一致，表明 1b 具有游离酶的亲和力，而无竞争抑制剂 2d 衍生物在变构部位具有很强的结合亲和力。与对映异构体 1b 相比，衍生自 d-缬氨酸的硫代海因 1a 的抑制、动力学和结合参数的立体化学影响非常大。
