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Title Site-directed mutagenesis of the membrane-proximal (MPER) α-helical E2 region of DENV E to characterise the significance of individual residues in protein expression and migration through the secretory pathway
Publication Date
Discipline/Department School of Chemistry and Molecular Biosciences
University/Publisher University of Queensland
Subjects/Keywords DENV-2; Dengue fever; Flavivirus; Virus Fusion; Structural Proteins; Envelope Proteins; 030406 Proteins and Peptides; 0601 Biochemistry and Cell Biology; 060108 Protein Trafficking
Language en
Country of Publication au
Record ID oai:espace.library.uq.edu.au:UQ:363952
Repository uqld1
Date Retrieved
Date Indexed 2020-08-18

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…63 63 64 68 69-70 70 83 99 108 12 Abbreviations: ADE bp C C6/36 CDC CO2 CPE CS DENV-2 DF DHF dH2O DI,II,III DMEM DMSO DNA DSS E EDTA eGFP ER FACS FCS FFWI FFWO g GAG GFP HCV HEK 293T HIV HS IF IRES JEV kB kDA LB M m.o.i. mAb MCS ml mM NC NCG nm…

…diseases, specifically of Dengue, Ross River fever and Malaria because the tropical zone has increased two degrees of latitude in the last 25 years [8]. In February 2011, there were 39 cases of DENV-2 in East Innisfail, QLD [9]. Figure…

…11). In DENV-2, E2 contains four phenylalanine and one tyrosine residues (figure 12 and 13). E1 and E2 are both amphipathic with one face positively charged, which may help alleviate electrostatic repulsion between the membrane-facing…

…fever), JE (Japanese encephalitis) and DEN (dengue). DEN refers to DENV-1 subtype. Helix 1 (E1) contains amino acids 398-420, helix 2 (E2) corresponds to residues 426448 [65]. F422G S L GG * VF429T…

…α-helical E2 domain of DENV E to obtain full-length prM and E mutant plasmids that will be used in all subsequent research (Chapter 3). 2. Develop a lentiviral expression system to assay fusion of these E2 mutants (Chapter 4). 3…

…cell prM and E expression, colocalisation, and cell surface trafficking (Chapter 6). 34 CHAPTER 2 MATERIALS AND METHODS 2.1. Vectors 2.1.1. pCI_NGCprME_opt_neo plasmid Figure 19. Plasmid map of pCI_NGCprME_opt_neo (New Guinea DENV 2

…CHAPTER 2 MATERIALS and METHODS 2.1. Vectors 2.1.1. pCI_NGCprME_opt_neo 2.1.2. pSFV and pSFVprME 2.1.3. Lentiviral Plasmids: pREV, pRRE, pMD.G, pLL3.7 pLL3.7, pGL-3, and pLL3.7_luc+ 2.2. Prokaryotic cell culture 2.2.1. Culture 2.2.2…

…pseudotyped viral particles and transduction of cells to test viral particles with VSV-G or DENV prME 4.3. Results: Luciferase reporter system 4.3.1. Luciferase plasmid cloning 4.3.2. Luciferase substrate assay 4.4. Discussion 4.4.1. Conclusions of lentiviral…