Thesis (Ph. D.)--University of Hong Kong, 1994. / Includes bibliographical references (leaves 254-266).

Thesis (Ph. D.)--University of Hong Kong, 2009. / Includes bibliographical references (leaves 131-151) Also available online.

Thesis (M.S.)--University of Wisconsin--Madison, 1970. / eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Influenza virus H1N1 is a huge threat on human health. Influenza occurs with seasonal variations and reaches peak prevalence in winter, with many people killed worldwide every year. In the research of interaction between influenza virus and cells, four major parts were in the range of our consideration, namely the proteins of virus, the proteome of host cell, the method of proteomic and the potencial medicine related with those significant proteins. Hemagglutinin (HA), as an envelope protein, plays an important role in influenza A virus. It was found that HA has a series of isoforms in two dimensional gels in this study. For the investigation of HA, firstly, virus was purified by sucrose density-gradient centrifugation, followed by the separation of virus proteins through electrophoresis method, and then these proteins were digested by different enzymes and analyzed through MALDI-TOF MS and ESI-Q-TOF MS. Database searching was used for identification of sequences. The results of the virus samples digested by different enzymes were compared, and the isoforms of HA were proved to be related with the glycan and their glycosylation sites. A novel strategy of stable-isotope N-phosphorylation labeling was developed for peptide de novo sequencing and protein quantification based on organic phosphorus chemistry. Different from other stable-isotope labeling reagents that needed to be activated in advance for peptide coupling, N-phosphorylation labeling reagents were activated in situ to form labeling intermediates with high activity and selectivity targeting on N-terminus and -amino group of lysine under various reaction conditions. The obtained results showed excellent correlation of the measured ratios to theoretical ratios with errors that ranging from 0.5 to 6.7 % and relative standard deviation of less than 10.6 %, indicating the reproducibility and preciseness of the developed method. The method development based on organic phosphorus chemistry offered a new approach for quantitative proteomics by using novel stable-isotope labeling reagents. A method combining hydrazide chemistry, stable isotope labeling and mass spectrometry analysis was developed and applied to study glycoproteins of H1N1 (A/Purto Rico/8/1934) infected cell line (A549). The result showed that some glycoproteins were significant in influenza virus infected cells. In these glycoproteins, RPC1_HUMAN, RHG25_HUMAN , RPTOR_HUMAN, ARHGC_HUMAN, ROCK1_HUMAN, DOCK3_HUMAN were down-regulated. Protein named TITIN_HUMAN, DESP_HUMAN, PTN13_HUMAN were up-regulated. High dose of N-acetylcysteine (NAC) was recently reported for a therapy of H1N1 influenza pneumonia. NAC was used as a small-molecule organic probe to investigate the protein expression of human lung carcinoma cell line (A549) infected by influenza virus H1N1. The obtained results showed that NAC kept cells away from apoptosis. Virus-infected cells were arrested in G0/G1 phase. The lowest cell population of G0/G1 phase was detected when the cells were treated by 10 mM NAC for one day. Software analysis showed that 4 proteins had close relationship. The results indicated that NAC as a small-molecule probe might effect the proteins expression of A549 cells infected by the H1N1 virus

No description available.

published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

Influenza infections continue to be a global health concern that causing both seasonal epidemics and unpredictable pandemics. Hemagglutinin (HA) and Neuraminidase (NA) are the two major surface glycoproteins of influenza viruses, which are important for their host cell sialic acid (Sia) receptor binding and cleaving activities. Although numerous methods have been developed to study the HA and NA interactions with sialic acid, x-ray crystallography remained the only method to provide detailed information at atomic resolution. The aim of this study is to develop and evaluate a novel strategy for the investigation of influenza virus-receptor interactions, which is able to provide information about an interaction down to atomic resolution. Influenza virus-like particles (VLPs) containing HA and NA separately were developed and it was reported here for the first time that sole expression of NA in mammalian cell led to VLP formation. Characterization of these VLPs demonstrated that they are non-infectious, but morphologically and biochemically mimic the native viruses. Therefore the VLPs can be regarded as an ideal research model to study the HA-Sia interaction without the interference of NA, or vice versa. Saturation transfer difference (STD) NMR spectroscopy is a state-of-the-art technology to determine how a binding-ligand interacts with its target protein. Modification of STD-NMR methodology was performed to adapt the technique to influenza VLP system. HA-Sia interaction was investigated in great detail and group epitope mapping of the interacting ligands was performed by analyzing the STD-NMR spectra. The data obtained are in a good agreement with the well established crystallography technique, reflecting the reliability of the STD-NMR technology. Regarding the NA-Sia interaction, my data demonstrated that substrate-hydrolysis specificity of NA is dependent on the binding of NA to those ligands. In addition, using competition experiments with NA inhibitor, a secondary sialic acid binding site was detected. It is the first direct experimental evidence that confirms avian, seasonal human and human pandemic swine-origin influenza virus N1 neuraminidases exhibit a distinct secondary binding site. In conclusion, here I presented a novel interdisciplinary strategy using VLP and NMR technology to study the interaction of influenza virus with its receptor. This method is unique in its ability to provide detailed information on the HA and NA interactions with sialic acid leading to group epitope mapping of the binding ligands, which will help us not only to understand the virus tropism but also to define new therapeutic targets. / published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

Thesis (Ph.D.)--Baylor University, 2008. / In abstract the '2' and '-/-' in NOD-SCID-[beta]2m-/- is superscript. In abstract the '+' after CD34 and CD8 is superscript. In abstract the '-' and '+' in CD45RA-CD27+CD4+ are superscript. Includes bibliographical references (p. 103-123).

Thesis (Ph. D.)--Florida State University, 2005. / Advisor: Dr. Timothy A. Cross, Florida State University, College of Arts and Sciences, Dept. of Chemistry and Biochemistry. Title and description from dissertation home page (viewed June 13, 2005). Document formatted into pages; contains xiv,160 pages. Includes bibliographical references.