The functional study of influenza B nucleoprotein.

Lam, Ka Han.Thesis (M.Phil.)--Chinese University of Hong Kong, 2011.Includes bibliographical references (leaves 77-82).Abstracts in English and Chinese.Acknowledgement --- p.iiAbstract --- p.iii摘要 --- p.vContent --- p.viiList of Abbreviations and Symbols --- p.xiChapter Chapter 1 --- IntroductionChapter 1.1 --- Severity of influenza --- p.1Chapter 1.2 --- Introduction of influenza viruses --- p.3Chapter 1.2.1 --- Virion and genome structure --- p.4Chapter 1.2.2 --- The replication cycle of influenza viruses --- p.5Chapter 1.3 --- Influenza virus NP --- p.8Chapter 1.3.1 --- The importance of NP in RNP structure maintenance --- p.9Chapter 1.3.2 --- NP self oligomerization --- p.10Chapter 1.3.3 --- NP-RNA interaction --- p.12Chapter 1.3.4 --- NP and other interacting partners --- p.13Chapter 1.4 --- Aim of the project --- p.16Chapter Chapter 2 --- Materials and MethodsChapter 2.1 --- Biological materials --- p.18Chapter 2.2 --- Construction of NP mutants --- p.19Chapter 2.3 --- Luciferase assay --- p.22Chapter 2.4 --- Western blot --- p.23Chapter 2.5 --- Protein expression and purification --- p.23Chapter 2.6 --- Circular dichroism spectroscopy --- p.24Chapter 2.7 --- Static Light scattering --- p.24Chapter 2.8 --- Surface plasmon resonance --- p.25Chapter 2.9 --- Co-immunoprecipitation (co-IP) --- p.26Chapter Chapter 3 --- Identification of residues crucial for NPB oligomerization and ribonucleoprotein activityChapter 3.1 --- Introduction --- p.27Chapter 3.2 --- Result --- p.31Chapter 3.2.1 --- NPB mutants showed deficiency in overall transcription and replication activity --- p.31Chapter 3.2.2 --- Expression and purification of NP mutants with low RNP activity --- p.37Chapter 3.2.2.1 --- Expression of MBP-tagged NP variants --- p.37Chapter 3.2.2.2 --- Purification of MBP-tagged NP variants --- p.38Chapter 3.2.3 --- Secondary structures of NP variants were comparable t o wild type NP --- p.41Chapter 3.2.4 --- NP variants with low RNP activity were abnormal in oligomerization in vitro --- p.42Chapter 3.2.5 --- NP variants with low RNP activity were impaired in homo-oligomer formation in vivo --- p.45Chapter 3.2.6 --- Discussion --- p.47Chapter Chapter 4 --- Identification of residues crucial for NP 一 RNA interaction and ribonucleoprotein activityChapter 4.1 --- Introduction --- p.56Chapter 4.2 --- Result --- p.58Chapter 4.2.1 --- NPB mutants showed deficiency in overall transcription and replication activity --- p.58Chapter 4.2.2 --- Expression and purification of NP variants with low RNP activity --- p.62Chapter 4.2.3 --- Secondary structures of NP variants were comparable t o wild type NP --- p.63Chapter 4.2.4 --- NP variants with low RNP activity were abnormal in RNA binding --- p.64Chapter 4.3 --- Discussion --- p.68Chapter Chapter 5 --- Conclusion and future prospect --- p.73Copyright --- p.76References --- p.7