Rufino Arcanjo Daniel Dias, Meneses Bezerra Isadora Basílio, Comerma-Steffensen Simón Gabriel, Prat-Duran Judit, Moreira Daniel Carneiro, Vasconcelos Andreanne Gomes, Plácido Alexandra, Teixeira Cátia, Gomes Paula, Cardoso-Teixeira Ana Carolina, Coelho-de-Souza Andrelina Noronha, Bemquerer Marcelo, Christensen Birgitte Mønster, Wimmer Reinhard, Pereira de Oliveira Aldeidia, Ramos Ricardo Martins, Simonsen Ulf, de Souza de Almeida Leite José Roberto
Pulmonary and Cardiovascular Pharmacology, Department of Biomedicine, Aarhus University, Aarhus, Denmark; Department of Biophysics and Physiology, Federal University of Piaui, Ministro Petrônio Portella Campus, Teresina, Brazil.
Department of Biophysics and Physiology, Federal University of Piaui, Ministro Petrônio Portella Campus, Teresina, Brazil; Information Systems Research Laboratory, Department of Information, Environment, Health and Food Production, Federal Institute of Piaui, IFPI, Teresina, Brazil.
Eur J Pharmacol. 2025 Sep 5;1002:177857. doi: 10.1016/j.ejphar.2025.177857. Epub 2025 Jun 19.
Previous studies have reported the involvement of argininosuccinate synthase (AsS) as a putative target for the cardiovascular effects of proline-rich oligopeptides. The present study investigated the mechanisms underlying the vasorelaxant effect of BPP-BrachyNH in rat small mesenteric arteries and applied in silico molecular dynamics studies to explore the interaction between AsS and BPP-BrachyNH. Segments of male rat mesenteric arteries mounted in microvascular myographs were treated with 10 μg/mL lipopolysaccharides (LPS), and then, BPP-BrachyNH was cumulatively added (10 - 3 × 10 M) to noradrenalin-contracted (1-10 μM) preparations. Molecular Dynamics calculations were performed between AsS enzyme (RCSB Protein Data Bank ID: 2NZ2) as the target and both BPP-BrachyNH and L-citrulline (PubChem CID 9750 code) as ligands. Immunohistochemistry showed expression of AsS and endothelial nitric oxide (NO) synthase (eNOS) in mesenteric arteries and of inducible NO synthase (iNOS) in segments exposed to LPS. The vasorelaxant effect of BPP-BrachyNH was abolished in the presence of 100 μM L-NNA (L-N-nitroarginine) and 3 μM ODQ ([1H-[1,2,4]-oxadiazolo-[4,3-a] quinoxalin-1-one]), and attenuated in the presence of 10 μM 1400W (N-(3-(aminomethyl)-benzyl)-acetamidine) and 1 mM MDLA (α-methyl-D, L-aspartic acid), inhibitors of iNOS and AsS, respectively. The AsS_BPP-BrachyNH complex showed increased binding energy, inhibition constant, and the number of interactions with amino acids when compared with the AsS_L-citrulline complex. These results suggest that the positive interaction of BPP-BrachyNH with AsS leads to L-citrulline recycling and increases L-arginine bioavailability, thereby improving the mechanism for the vasorelaxant effect. Our findings open new perspectives for potential therapeutic applications of proline-rich oligopeptides in vascular dysfunction.
先前的研究报道，精氨酸琥珀酸合酶（AsS）可能是富含脯氨酸的寡肽产生心血管效应的作用靶点。本研究调查了BPP-BrachyNH对大鼠小肠系膜动脉血管舒张作用的潜在机制，并通过计算机模拟分子动力学研究来探索AsS与BPP-BrachyNH之间的相互作用。将雄性大鼠肠系膜动脉段安装在微血管肌动描记器上，用10μg/mL脂多糖（LPS）处理，然后将BPP-BrachyNH（10 - 3×10 M）累积添加到去甲肾上腺素收缩（1 - 10μM）的标本中。以AsS酶（RCSB蛋白质数据库ID：2NZ2）为靶点，以BPP-BrachyNH和L-瓜氨酸（PubChem CID 9750代码）为配体进行分子动力学计算。免疫组织化学显示，肠系膜动脉中存在AsS和内皮型一氧化氮合酶（eNOS）的表达，在暴露于LPS的节段中存在诱导型一氧化氮合酶（iNOS）的表达。在存在100μM L-NNA（L-N-硝基精氨酸）和3μM ODQ（[1H-[1,2,4]-恶二唑-[4,3-a]喹喔啉-1-酮]）的情况下，BPP-BrachyNH的血管舒张作用消失，而在分别存在10μM 1400W（N-(3-(氨甲基)-苄基)-乙脒）和1mM MDLA（α-甲基-D，L-天冬氨酸）（iNOS和AsS的抑制剂）的情况下，其血管舒张作用减弱。与AsS_L-瓜氨酸复合物相比，AsS_BPP-BrachyNH复合物显示出更高的结合能、抑制常数以及与氨基酸的相互作用数量。这些结果表明，BPP-BrachyNH与AsS的正向相互作用导致L-瓜氨酸循环利用并增加L-精氨酸的生物利用度，从而改善血管舒张作用的机制。我们的研究结果为富含脯氨酸的寡肽在血管功能障碍中的潜在治疗应用开辟了新的前景。
