ERA

Download the full-sized PDF of Development of a Spectral Searching Strategy for Peptide and Protein IdentificationDownload the full-sized PDF

Analytics

Share

Permanent link (DOI): https://doi.org/10.7939/R3NV99N4Z

Download

Export to: EndNote  |  Zotero  |  Mendeley

Communities

This file is in the following communities:

Graduate Studies and Research, Faculty of

Collections

This file is in the following collections:

Theses and Dissertations

Development of a Spectral Searching Strategy for Peptide and Protein Identification Open Access

Descriptions

Other title
Subject/Keyword
Proteomics
Spectral Library
Isotopic Labeling
Spectral Searching
Protein Identification
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Xu, Mingguo
Supervisor and department
Li, Liang (Chemistry)
Examining committee member and department
Campbell, Robert (Chemistry)
Lubman, David (Surgery)
Lin, Guohui (Computer Science)
Clive, Derrick (Chemistry)
Serpe, Michael (Chemistry)
Department
Department of Chemistry
Specialization

Date accepted
2012-08-20T12:54:17Z
Graduation date
2012-11
Degree
Doctor of Philosophy
Degree level
Doctoral
Abstract
The overall goal of this thesis research is to develop a spectral searching strategy capable of identifying peptide sequences from MS/MS spectra with high sensitivity and accuracy. First, a shotgun proteome analysis method was developed and successfully applied to the identification of proteins from thousands of cancer cells. This work illustrated that proteome profiling of a small number of cells isolated from blood can be achieved. By comparing the obtained profile to a standard profile, cell typing might also be possible. This method may prove to be useful for cancer diagnosis or prognosis. From this study, we realized that sequence database searching strategy is one of the bottlenecks to achieve better sensitivity of protein identification for proteome profiling work. As a promising alternative, spectral searching strategy is believed to be able to provide more sensitive and accurate peptide and protein identification. In spectral searching strategy, there are two main components: spectral libraries and the searching algorithm. Since an accurate identification by spectral searching strategy is built on the premise of a reliable MS/MS spectral library, 15N-metabolic labeling and 18O-labeling approaches were developed to experimentally validate all the peptide matches from sequence database search results. With those validated matches, the sensitivity and accuracy of commonly used search engines (Mascot and X!Tandem) and two popular statistical approaches (PeptideProphet and Percolator) were carefully examined. Moreover, two strategies were designed to identify single-hit protein identifications (proteins identified by only one peptide) with high reliability. In addition, Percolator was successfully interfaced with X!Tandem to enhance its performance. Finally, a spectral searching algorithm called SpecMatching was developed to utilize the experimentally validated spectral library. In analyzing a digest of an E. coli extract using both Mascot and SpecMatching, it was shown that SpecMatching provided better sensitivity and specificity even with this small-size spectral library.
Language
English
DOI
doi:10.7939/R3NV99N4Z
Rights
Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
Citation for previous publication

File Details

Date Uploaded
Date Modified
2014-05-01T00:57:29.659+00:00
Audit Status
Audits have not yet been run on this file.
Characterization
File format: pdf (Portable Document Format)
Mime type: application/pdf
File size: 3018086
Last modified: 2015:10:12 20:52:48-06:00
Filename: Xu_Mingguo_Fall 2012.pdf
Original checksum: 3fdb83a440dbfec0723fdb543923c23d
Well formed: true
Valid: true
File author: MX
Page count: 244
File language: en-CA
Activity of users you follow
User Activity Date