Microbial biofilms play a critical role in environmental biotechnology and associated applications. Biofilm production can be enhanced by inhibiting the function of proteins that negatively regulate their formation. With this objective, an in silico approach was adopted to identify competitive inhibitors of eight biofilm-antagonistic proteins, namely AbrB and SinR (from Bacillus subtilis) and AmrZ, PDE (EAL), PslG, RetS, ShrA and TpbA (from Pseudomonas aeruginosa). Fifteen inhibitors that structurally resembled the natural ligand of each protein were shortlisted using ligand-based and structure-based virtual screening. The top four inhibitors obtained from molecular docking using Autodock Vina were further docked using SwissDock and DOCK 6.9 to obtain a consensus hit for each protein based on different scoring functions. Further analysis of the protein–ligand complexes revealed that these top inhibitors formed significant non-covalent interactions with their respective protein binding sites. The eight protein-ligand complexes were then subjected to molecular dynamics simulations for 30 ns using GROMACS. RMSD and radius of gyration values of 0.1–0.4 nm and 1.0–3.5 nm, respectively, along with hydrogen bond formation throughout the trajectory indicated that all the complexes remained stable, compact and intact during the simulation period. Binding energy values between –20 and –77 kJ/mol obtained from MM-PBSA calculations further confirmed the high affinities of the eight inhibitors for their respective receptors. The outcome of this study holds great promise to enhance biofilms that are central to biotechnological processes associated with microbial electrochemical technologies, wastewater treatment, bioremediation and the industrial production of value-added products.

Graphic abstract

中文翻译：

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识别生物膜拮抗蛋白的潜在抑制剂以促进生物膜形成：虚拟筛选和分子动力学模拟方法

微生物生物膜在环境生物技术和相关应用中发挥着关键作用。可以通过抑制负调控其形成的蛋白质的功能来增强生物膜的产生。为此，采用计算机模拟方法来鉴定八种生物膜拮抗蛋白的竞争性抑制剂，即 AbrB 和 SinR（来自枯草芽孢杆菌）和 AmrZ、PDE (EAL)、PslG、RetS、ShrA 和 TpbA（来自铜绿假单胞菌））。使用基于配体和基于结构的虚拟筛选将 15 种结构类似于每种蛋白质的天然配体的抑制剂入围。使用 Autodock Vina 从分子对接获得的前四种抑制剂使用 SwissDock 和 DOCK 6.9 进一步对接，以获得基于不同评分函数的每种蛋白质的一致命中。对蛋白质-配体复合物的进一步分析表明，这些顶级抑制剂与其各自的蛋白质结合位点形成了显着的非共价相互作用。然后使用 GROMACS 对八种蛋白质-配体复合物进行 30 ns 的分子动力学模拟。RMSD 和回转半径值分别为 0.1-0.4 nm 和 1.0-3.5 nm，随着整个轨迹的氢键形成表明所有配合物都保持稳定，在模拟期间紧凑且完整。从 MM-PBSA 计算中获得的介于 –20 和 –77 kJ/mol 之间的结合能值进一步证实了八种抑制剂对其各自受体的高亲和力。这项研究的结果很有希望增强生物膜，这些生物膜对于与微生物电化学技术、废水处理、生物修复和增值产品的工业生产相关的生物技术过程至关重要。

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