Department of Biotechnology, Institute of Science and High Technology and Environmental Sciences, Graduate University of Advanced Technology, Kerman, Iran.
Microbiology Department, Burn and Wound Healing Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.
Microbiol Spectr. 2023 Aug 17;11(4):e0463022. doi: 10.1128/spectrum.04630-22. Epub 2023 Jul 12.
Circulating influenza A virus provided an excellent opportunity to study the adaptation of the influenza A(H1N1)pdm09 virus to the human host. Particularly, due to the availability of sequences taken from isolates, we could monitor amino acid changes and the stability of mutations that occurred in hemagglutinin (HA). HA is crucial to viral infection because it binds to ciliated cell receptors and mediates the fusion of cells and viral membranes; because antibodies that bind to HA may block virus entry to the cell, this protein is subjected to high selective pressure. In this study, the locations of mutations in the structures of mutant HA were analyzed and the three-dimensional (3D) structures of these mutations were modeled in I-TASSER. Also, the location of these mutations was visualized and studied using Swiss PDB Viewer software and the PyMOL Molecular Graphics System. The crystal structure of the HA from A/California/07/2009 (3LZG) was used for further analysis. The new noncovalent bond formations in mutant luciferases were analyzed via WHAT IF and PIC, and protein stability was evaluated in the iStable server. We identified 33 and 23 mutations in A/Shiraz/106/2015 and A/California/07/2009 isolates, respectively; some mutations are located on the antigenic sites of Sa, Sb, Ca1, Ca2, and Cb HA1 and the fusion peptide of HA2. The results show that with the mutation some interactions are lost and new interactions are formed with other amino acids. The results of the free-energy analysis suggested that these new interactions have a destabilizing effect, which needs confirmation experimentally. Due to the fact that the mutations that occurred in the influenza virus HA cause the instability of the protein produced by the virus and antigenic changes and the escape of the virus from the immune system, the mutations that occurred in A/Shiraz/1/2013 were investigated in terms of energy level and stability. The mutations located in a globular portion of the HA are S188T, Q191H, S270P, K285Q, and P299L. On the other hand, the E374K, E46K-B, S124N-B, and I321V mutations are located in the stem portion of the HA (HA2). The change V252L mutation eliminates interactions with Ala181, Phe147, Leu151, and Trp153 and forms new interactions with Gly195, Asn264, Phe161, Met244, Tyr246, Leu165, and Trp167 which can change the stability of the HA structure. The K166Q mutation, which is located within the antigenic site Sa, causes the virus to escape from the immune response.
循环流感病毒为研究流感 A(H1N1)pdm09 病毒对人类宿主的适应性提供了极好的机会。特别是由于可以从分离物中获取序列，我们可以监测到血凝素 (HA) 中发生的氨基酸变化和突变的稳定性。HA 对病毒感染至关重要，因为它与纤毛细胞受体结合并介导细胞和病毒膜的融合；因为与 HA 结合的抗体可能阻止病毒进入细胞，所以该蛋白受到高度的选择性压力。在这项研究中，分析了突变 HA 结构中的突变位置，并使用 I-TASSER 对这些突变的三维 (3D) 结构进行了建模。此外，还使用 Swiss PDB Viewer 软件和 PyMOL 分子图形系统对这些突变的位置进行了可视化和研究。使用 A/California/07/2009（3LZG）的 HA 晶体结构进行进一步分析。通过 WHAT IF 和 PIC 分析了突变荧光素中的新非共价键形成，并在 iStable 服务器中评估了蛋白质稳定性。我们分别在 A/Shiraz/106/2015 和 A/California/07/2009 分离株中鉴定出 33 和 23 个突变；有些突变位于 Sa、Sb、Ca1、Ca2 和 Cb HA1 和 HA2 融合肽的抗原位点上。结果表明，随着突变的发生，一些相互作用丢失了，与其他氨基酸形成了新的相互作用。自由能分析的结果表明，这些新的相互作用具有不稳定的作用，这需要实验证实。由于流感病毒 HA 中的突变导致病毒产生的蛋白质不稳定和抗原变化以及病毒逃避免疫系统，因此研究了 A/Shiraz/1/2013 中发生的突变的能量水平和稳定性。位于 HA 球形部分的突变是 S188T、Q191H、S270P、K285Q 和 P299L。另一方面，E374K、E46K-B、S124N-B 和 I321V 突变位于 HA（HA2）的茎部。V252L 突变消除了与 Ala181、Phe147、Leu151 和 Trp153 的相互作用，并与 Gly195、Asn264、Phe161、Met244、Tyr246、Leu165 和 Trp167 形成新的相互作用，这可能改变 HA 结构的稳定性。位于抗原位点 Sa 内的 K166Q 突变导致病毒逃避免疫反应。
