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Graduate School of Ajou University Department of Molecular Science and Technology 4. Theses(Ph.D)

Structural insights into the immune subversion of coronaviruses and identification of potent small molecule inhibitors using computational approaches

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dc.contributor.advisorSangdun Choi-
dc.contributor.authorBATOOL MARIA-
dc.date.accessioned2022-11-29T02:32:14Z-
dc.date.available2022-11-29T02:32:14Z-
dc.date.issued2020-08-
dc.identifier.other30243-
dc.identifier.urihttps://dspace.ajou.ac.kr/handle/2018.oak/19739-
dc.description학위논문(박사)--아주대학교 일반대학원 :분자과학기술학과,2020. 8-
dc.description.tableofcontentsCHAPTER 1 1 Introduction 1 1.1 Overview of Coronaviruses 2 1.2 Entry and replication of viruses 3 1.3 Potential therapeutic targets 6 CHAPTER 2 9 Methods 9 2.1 Protein modeling and interface analysis 10 2.2 Alanine scanning mutagenesis 10 2.3 Virtual screening and molecular docking 11 2.4 Molecular dynamic simulations 12 2.5 Principal component analysis 15 2.6 Free energy landscape 15 2.7 MMPBSA 16 2.8 Data analysis and visualization 17 CHAPTER 3 18 Results and Discussion 1 18 Structural insights into the Middle East respiratory syndrome coronavirus 4a protein and its dsRNA binding mechanism 19 3.1 Summary 20 3.2 3D modeling of p4a-dsRNA complex and interface analyses 20 3.3 Identification of hotspot residues and their influence on p4a-dsRNA binding mechanism 23 3.4 Investigation of impact of hotspot residues on p4a-dsRNA complex through structural dynamics 24 3.5 Interactions analysis between p4a-dsRNA wild type complex and mutants 27 3.6 Dynamic motions of MERS-CoV p4a mutants 27 3.7 Transition of proteins from unfolded to native state 31 3.8 Binding free energy calculation 32 3.9 Discussion 33 CHAPTER 4 36 Results and Discussion 2 36 Discovery of potential inhibitors to block MERS-CoV evasion from the host immune system by targeting a non-structural accessory protein 37 4.1 Summary 38 4.2 Understanding of dynamic behavior and stability of p4a 38 4.3 Identification of potential compound to block p4a-dsRNA interaction 39 4.4 Identification of physicochemical and pharmacokinetic properties 45 4.5 Discussion 46 CHAPTER 5 48 Results and Discussion 3 48 Structural insights into the behavior of the catalytic site of SARS-CoV-2 main protease 49 5.1 Summary 50 5.2 Analysis of apo-3CL protease behavior via MD simulations 50 5.3 Dynamic behavior of 3CL protease with different ligands 52 5.4 The behavior of 3CL protease catalytic site in the presence of ligands 58 5.5 The binding energy of ligands toward 3CL protease 60 5.6 Discussion 61 Bibliography 64 Appendix 1 74 Appendix 2 75 Appendix 3 76-
dc.language.isoeng-
dc.publisherThe Graduate School, Ajou University-
dc.rights아주대학교 논문은 저작권에 의해 보호받습니다.-
dc.titleStructural insights into the immune subversion of coronaviruses and identification of potent small molecule inhibitors using computational approaches-
dc.typeThesis-
dc.contributor.affiliation아주대학교 일반대학원-
dc.contributor.alternativeNameBATOOL MARIA-
dc.contributor.department일반대학원 분자과학기술학과-
dc.date.awarded2020. 8-
dc.description.degreeDoctoral-
dc.identifier.localId1151650-
dc.identifier.uciI804:41038-000000030243-
dc.identifier.urlhttp://dcoll.ajou.ac.kr:9080/dcollection/common/orgView/000000030243-
dc.subject.keywordCorona viruses-
dc.subject.keywordMD Simulations-
dc.subject.keywordVirtual screening-
dc.subject.keywordcomputational biology-
dc.description.alternativeAbstractThe recent occurrence of COVID-19 pandemic causing fever, respiratory illness, and pneumonia, has drawn the attention across the globe and has been declared a global health emergency. Among humans, coronaviruses were not believed to be pathogenic until the severe acute respiratory syndrome (SARS) epidemic occurred in late 2002. Ten years later in 2012, a new coronavirus epidemic named as Middle East respiratory syndrome (MERS) emerged in Asian and Middle Eastern countries. Several studies have been published to understand the cell entry, replication, and protein structures of virus. The genomic organization of SARS and MERS is similar, comprising an enveloped, positive-sense, single-stranded RNA genome that translates into various structural and non-structural proteins (nsps). After viral entry into the host cell, virus releases its RNA into the cytoplasm where it is translated into polyproteins pp1a and pp1ab. These polyproteins are cleaved by proteases that produce 16 nsps. The viral genome is translated, replicated, and assembled in the cytoplasm which is then released via exocytosis. To combat with the coronavirus diseases, it is important to block viral infection or replication. Therefore, two potential target proteins were selected for the drug development. The first target is an accessory protein 4a of MERS-CoV which has been proved through in vitro and in vivo studies that it is involved in inhibition of IFN production which influences the viral pathogenicity. 3D structure of protein 4a was modeled followed by structural validation and computational alanine mutagenesis. Residues N8, K27, W45, K63, and K67 were selected based on the dStability and dAffinity values. The mutant and wild type complexes were simulated for 200 ns each. The MD simulation results revealed that K27, W45, K63, and K67 are key residues that play crucial role in RNA binding. Thus, suggesting that blocking these residual interactions could destabilize the 4a-RNA complex. Furthermore, considering these residues a part of binding site, virtual screening of antiviral as well as druglike and leadlike compounds from ZINC database is conducted using the most stable confirmation extracted from MD simulations. The ten compounds from both datasets were selected based on binding affinity scores and their binding pose. The interactions were analyzed in detail along with the free binding energy calculations. Most of the selected compounds possess antiviral properties. These compounds can be tested through in vitro and in vivo experiments. The 3CL protease is another potential target due to its proteolytic activity that cleaves the polyprotein. Many drugs have been repositioned, and a few ligands are cocrystalized with SARS-CoV-2 3CL protease. The flexibility of the catalytic site leads to the conformational changes that influences the ligand binding. Thus, it is necessary to probe the flexibility and rigidity of protein’s active site. 100 ns MD simulations were performed for apo form of 3CL protease and its complexes with different cocrystalized ligands. The trajectory analysis and binding free energy calculation revealed that the ligand that can break the catalytic dyad bond in the active site would be a potential candidate to block the proteolytic activity. The alpha-keto group present in the inhibitor of 6LU7 make strong bonds with both residues Cys and His of catalytic dyad. Therefore, addition of such chemical groups that break the bond between Cys and His would be important to block the functional 3CL protease. Conclusively, both target proteins 4a and 3CL protease are important for viral pathogenicity and replication, respectively. Therefore, understanding of structure dynamics and insights into the binding site is crucial for development of an efficient drug molecule. 3CL protease is structurally and functionally conserved protein among all three coronaviruses. Thus, targeting this protein would be a hope to combat all coronavirus diseases.-
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