Catalysts and Organic Synthesis Research Laboratory, Department of Chemistry, Iran University of Science and Technology, Tehran 16846-13114, Iran.
Catalysts and Organic Synthesis Research Laboratory, Department of Chemistry, Iran University of Science and Technology, Tehran 16846-13114, Iran.
Ultrason Sonochem. 2020 Mar;61:104824. doi: 10.1016/j.ultsonch.2019.104824. Epub 2019 Oct 8.
Herein, a novel heterogeneous nanoscale reducing agent for antibody cleavage, made of iron oxide nanoparticles, silica network, palladium on calcium carbonate (10%), and dithiothreitol (FeO@Pd/CaCO-DTT), is presented as a substantial alternative for traditional homogeneous analogues. Conventionally, antibody fragmentation is accomplished using reducing agents and proteases that digest or cleave certain portions of the immunoglobulin protein structure to provide active thiol sites for drug tagging aims. Then, dialysis process is needed to separate excess chemical structures and purify the reduced antibody. In this work, we have made an effort to design a suitable heterogeneous tool for protein cleavage and skip the dialysis process for purification of the reduced antibody. In this regard, firstly, various preparation methods including microwave irradiation, reflux and ultrasonication have been precisely compared, and it has been proven that the best result is obtained through 10 min ultrasound (US) irradiation using an US bath with 50 KHz frequency and 200 W L power density. Then, all the necessary structural analyses have been done and thoroughly interpreted for the final product. Afterward, the catalytic performance of FeO@Pd/CaCO-DTT nanoscale system in the presence of US waves (50 KHz, 200 W) has been monitored using some disulphide derivatives. The NPs could be conveniently separated from the mixture through their substantial paramagnetic property. Thus, dialysis process in which various types of membranes are used is practically jumped after the reduction process. In this work, this is clearly demonstrated that there is a constructive synergistic effect between US waves and prepared FeO@Pd/CaCO-DTT nanoscale reducing agent. Ultimately, trastuzumab (anti HER-2) antibody has been used to test the performance of the prepared FeO@Pd/CaCO-DTT NPs in a real protein reduction reaction.
本文提出了一种新型的用于抗体切割的异相纳米还原剂，由氧化铁纳米粒子、硅网络、碳酸钙负载的钯（10%）和二硫苏糖醇（FeO@Pd/CaCO-DTT）组成，是传统均相类似物的一种实质性替代物。传统上，使用还原剂和蛋白酶来完成抗体片段化，这些还原剂和蛋白酶可以消化或切割免疫球蛋白蛋白质结构的某些部分，为药物标记目的提供活性硫醇位点。然后，需要通过透析过程来分离多余的化学结构并纯化还原的抗体。在这项工作中，我们努力设计了一种合适的用于蛋白质切割的异相工具，并跳过透析过程来纯化还原的抗体。在这方面，首先，精确比较了包括微波辐射、回流和超声在内的各种制备方法，结果证明通过在 50 KHz 频率和 200 W·L 功率密度的超声浴中使用 10 分钟超声（US）辐射，可以获得最佳结果。然后，对最终产物进行了所有必要的结构分析，并进行了详细解释。随后，在 US 波（50 KHz，200 W）的存在下，监测了 FeO@Pd/CaCO-DTT 纳米系统的催化性能。可以通过 NPs 的强顺磁性方便地将它们从混合物中分离出来。因此，在还原过程之后，实际上可以跳过使用各种类型的膜的透析过程。在这项工作中，清楚地证明了 US 波和制备的 FeO@Pd/CaCO-DTT 纳米还原剂之间存在建设性的协同效应。最终，使用曲妥珠单抗（抗 HER-2）抗体来测试制备的 FeO@Pd/CaCO-DTT NPs 在实际蛋白质还原反应中的性能。
