Malaria and Vector Research Group, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
Chemical and Petroleum Engineering Department (BBRC), Sharif University of Technology, Tehran, Iran.
PLoS One. 2022 Sep 12;17(9):e0274275. doi: 10.1371/journal.pone.0274275. eCollection 2022.
In order to move towards the elimination and eradication of malaria in the world, the development of vaccines is inevitable. Many modern vaccines are based on recombinant technology; however, they may not provide a fully protective, long-lasting immune response. One of the strategies to improve recombinant vaccines is designing the nanovaccines such as self-assembling protein nanoparticles (SAPNs). Hence, the presentation of epitopes in a repeat array and correct conformation should be considered. P. falciparum generative cell-specific 1 (PfGCS1) is a main transmission-blocking vaccine candidate with two highly conserved fragments, HAP2-GCS1 and cd loop, inducing partial malaria transmission inhibitory antibodies. Therefore, to design an effective malaria vaccine, we used cd loop and HAP2-GCS1 fragments at the amino and carboxy terminuses of the SAPN-forming amino acid sequence, respectively.
METHODOLOGY/PRINCIPAL FINDINGS: The SAPN monomer (PfGCS1-SAPN) sequence was designed, and the three-dimensional (3D) structure was predicted. The result of this prediction ensured the presence of antigens on the SAPN surface. Then the accuracy of the predicted 3D structure and its stability were confirmed by 100 ns molecular dynamics (MD) simulation. The designed SAPN substructure sequence was synthesized, cloned, and expressed in Escherichia coli. With a gradual decrease in urea concentration in dialysis solutions, the purified proteins progressed to the final desired structure of the SAPN, which then was confirmed by Dynamic Light Scattering (DLS) and Field Emission Scanning Electron Microscopy (FESEM) tests. According to the Enzyme-Linked Immunosorbent Assay (ELISA), antigenic determinants were presented on the SAPN surface and interacted with antibodies in the serum of malaria patients.
CONCLUSIONS/SIGNIFICANCE: Our results show that the SAPN formed by PfGCS1-SAPN has produced the correct shape and size, and the antigenic determinants are presented on the surface of the SAPN, which indicates that the designed SAPN has great potential to be used in the future as a malaria vaccine.
为了在全球范围内消除和消灭疟疾，疫苗的开发是必不可少的。许多现代疫苗都是基于重组技术的；然而，它们可能无法提供完全保护、持久的免疫反应。提高重组疫苗的一种策略是设计纳米疫苗，如自组装蛋白纳米颗粒 (SAPN)。因此，应该考虑在重复阵列中呈现表位和正确的构象。恶性疟原虫生殖细胞特异性 1 (PfGCS1) 是一种主要的阻断传播疫苗候选物，具有两个高度保守的片段，HAP2-GCS1 和 cd 环，诱导部分疟疾传播抑制抗体。因此，为了设计有效的疟疾疫苗，我们分别在 SAPN 形成氨基酸序列的氨基和羧基末端使用 cd 环和 HAP2-GCS1 片段。
方法/主要发现：设计了 SAPN 单体（PfGCS1-SAPN）序列，并预测了其三维（3D）结构。该预测结果确保了 SAPN 表面存在抗原。然后，通过 100ns 分子动力学 (MD) 模拟确认预测的 3D 结构的准确性及其稳定性。合成、克隆并在大肠杆菌中表达设计的 SAPN 亚结构序列。随着透析液中尿素浓度的逐渐降低，纯化的蛋白质逐渐形成 SAPN 的最终所需结构，然后通过动态光散射 (DLS) 和场发射扫描电子显微镜 (FESEM) 测试进行确认。根据酶联免疫吸附测定 (ELISA)，SAPN 表面呈现抗原决定簇并与疟疾患者血清中的抗体相互作用。
结论/意义：我们的结果表明，由 PfGCS1-SAPN 形成的 SAPN 产生了正确的形状和大小，并且抗原决定簇呈现在 SAPN 的表面上，这表明设计的 SAPN 具有很大的潜力，可在未来用作疟疾疫苗。
