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Identification of Protein Isoforms i...

Chang, Kung-yen.

Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases.

紀錄類型:	書目-電子資源 : Monograph/item
正題名/作者:	Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases.
作者:	Chang, Kung-yen.
面頁冊數:	104 p.
附註:	Source: Dissertation Abstracts International, Volume: 72-10, Section: B, page: 5656.
附註:	Adviser: David C. Muddiman.
Contained By:	Dissertation Abstracts International72-10B.
標題:	Chemistry, Analytical.
電子資源:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3463755
ISBN:	9781124753065

Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases.
Chang, Kung-yen.

Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases. - 104 p.

Source: Dissertation Abstracts International, Volume: 72-10, Section: B, page: 5656.

Thesis (Ph.D.)--North Carolina State University, 2011.

Tandem mass spectrometry has been one of the most effective high-throughput approaches for protein identification and quantification. Database searching is the most frequently used approach for automated peptide assignment of tandem mass spectra. Protein identifications are inferred by grouping the peptide-spectrum matches. However, MS/MS search engines depend on protein databases to provide candidates for consideration. Alternative splicing of precursor messenger RNA expands the diversity of eukaryotic proteome in different tissues and developmental stages by allowing individual genes to generate multiple protein isoforms with different functions. Aberrant splicing has been implicated in various diseases including cancer. Many protein databases only include a limited number of isoforms to keep minimal redundancy. As a result, the database search will not be able to identify the true protein isoforms even with high quality tandem MS data and accurate intact precursor ion mass. The primary goal of the studies described in this dissertation is to investigate whether total protein identifications will increase after introducing the eligible splice isoforms for database searching. The related topics including the performance of search algorithms, the evaluation of alignment programs, and temporal profiles of potential isoforms are investigated as well.

ISBN: 9781124753065Subjects--Topical Terms:

224793
Chemistry, Analytical.

Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases.
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245 1 0 $a Identification of Protein Isoforms in Mass Spectrometry Based Proteomic Analyses Using Alternative Splicing Databases.
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500 $a Source: Dissertation Abstracts International, Volume: 72-10, Section: B, page: 5656.
500 $a Adviser: David C. Muddiman.
502 $a Thesis (Ph.D.)--North Carolina State University, 2011.
520 $a Tandem mass spectrometry has been one of the most effective high-throughput approaches for protein identification and quantification. Database searching is the most frequently used approach for automated peptide assignment of tandem mass spectra. Protein identifications are inferred by grouping the peptide-spectrum matches. However, MS/MS search engines depend on protein databases to provide candidates for consideration. Alternative splicing of precursor messenger RNA expands the diversity of eukaryotic proteome in different tissues and developmental stages by allowing individual genes to generate multiple protein isoforms with different functions. Aberrant splicing has been implicated in various diseases including cancer. Many protein databases only include a limited number of isoforms to keep minimal redundancy. As a result, the database search will not be able to identify the true protein isoforms even with high quality tandem MS data and accurate intact precursor ion mass. The primary goal of the studies described in this dissertation is to investigate whether total protein identifications will increase after introducing the eligible splice isoforms for database searching. The related topics including the performance of search algorithms, the evaluation of alignment programs, and temporal profiles of potential isoforms are investigated as well.
520 $a Dozens more tandem mass spectra were assigned to the peptides in alternative splicing databases and putative isoforms were detected in two fungi, Aspergillus flavus and Magnaporthe oryzae. Sometimes, MS/MS spectra alone were insufficient to confirm the discoveries of splice isoforms. Part of the discoveries could be contributed by annotation errors. The findings of two isoform proteins having different functional domains were reported in both organisms. The comparison of search engines showed large variations between the splice variants identified by different algorithms. The consensus peptides from multiple algorithms were used to select the top candidates for valid identifications. The alternative splicing databases generated from different alignment programs varied in size but the majority of predicted sequences were identical. Similar lists of putative isoforms were found in M. oryzae spores regardless of what alignment program was used to produce the alternative splicing database. The consistency was probably derived from the rational constraints applied during construction. In the temporal analysis of spore development induced by cAMP in M. oryzae, the expression profile of one putative isoform altered while the other protein seemed unaffected.
520 $a Current genome annotations could contain errors and major protein databases are incomplete. Alternatively spliced isoforms need to be considered while make the inference of protein identification by tandem mass spectrometry. The introduction of an alternative splicing database helped identify more proteins and unveiled more information about the proteome. In addition to the discovery of splice variants, AS database also showed potential to improve genome annotation. The increase of available mRNAs/ESTs improved the quality of the alternative splicing database. The multiple search engines approach can be used to validate the quality of the protein identifications. The variance between alignment programs can be reduced and the most significant isoforms can be captured through applying appropriate constraints in the prediction pipeline.
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