Department of Pharmaceutical Biotechnology, School of Pharmacy and Pharmaceutical Sciences, Isfahan University of Medical Sciences, Isfahan, The Islamic Republic of Iran.
Faculty of Science, Department of Biology, Shahrekord University, Shahrekord, The Islamic Republic of Iran.
J Biomol Struct Dyn. 2021 Mar;39(4):1321-1333. doi: 10.1080/07391102.2020.1729865. Epub 2020 Mar 2.
Plasminogen activators (PAs) are widely used for treatment of disorders caused by clot formation. Fibrin specific PAs are safe drugs from this group because of reducing the incidence of hemorrhage. The newer generation of PAs like tenecteplase, reteplase and desmoteplase were designed with the aim of achieving desirable properties such as improving specificity and affinity to fibrin and increasing half-life. Protein engineering and using of theoretical methods can help to rational and reliable design of new PAs with a set of favorable properties. In the present study, two new chimeric reteplase named M1-chr and M2-chr were designed with the aim of enhancing fibrin affinity also some potential properties include of increasing resistance to plasminogen activator inhibitor-1 and decreasing neurotoxicity. So, finger domain of desmoteplase was added to reteplase as a high fibrin specific domain. Some other point mutations were considering to achieve other mentioned properties. Three dimensional structure of wild-type reteplase and mutants were created by homology modeling and were evaluated by molecular dynamic simulation. Then, mutants docked to fibrin by HADDOCK web tools. Result of theoretical section verified the stability of mutants' structures. Also showed better interaction between M1-chr with fibrin than M2-chr. Wild-type and mutants were produced in bacterial expression system. Experimental assessment showed both mutants have appropriate enzymatic activity also 1.9-fold fibrin binding ability compared to wild-type. Therefore, this study offers new thrombolytic drugs with desirable properties specially enhanced fibrin affinity so they can represent a promising future in cost-effective production of favorable thrombolytic drugs.Communicated by Ramaswamy H. Sarma.
纤溶酶原激活剂 (PAs) 广泛用于治疗由血栓形成引起的疾病。纤维蛋白特异性 PA 是该类药物中较为安全的药物，因为其可降低出血的发生率。新一代的 PA，如替奈普酶、瑞替普酶和地西普酶，旨在实现改善纤维蛋白特异性和亲和力、增加半衰期等理想特性。蛋白质工程和理论方法的使用有助于合理可靠地设计具有一系列优良特性的新型 PA。在本研究中，设计了两种新的嵌合瑞替普酶，分别命名为 M1-chr 和 M2-chr，旨在提高纤维蛋白亲和力，同时还具有一些潜在的特性，包括增加对纤溶酶原激活物抑制剂-1 的抗性和降低神经毒性。因此，将地西普酶的指状结构域添加到瑞替普酶中作为高纤维蛋白特异性结构域。还考虑了一些点突变来实现其他提到的特性。通过同源建模创建了野生型瑞替普酶和突变体的三维结构，并通过分子动力学模拟进行了评估。然后，通过 HADDOCK web 工具将突变体对接至纤维蛋白。理论部分的结果验证了突变体结构的稳定性。同时表明 M1-chr 与纤维蛋白的相互作用优于 M2-chr。野生型和突变体在细菌表达系统中进行了生产。实验评估表明，与野生型相比，两种突变体都具有适当的酶活性和 1.9 倍的纤维蛋白结合能力。因此，本研究提供了具有理想特性的新型溶栓药物，特别是增强了纤维蛋白亲和力，因此它们在具有成本效益的生产有前途的溶栓药物方面具有广阔的前景。由 Ramaswamy H. Sarma 交流。
