Chemical labelling methods for gel free based proteomics
(2007)- Abstract
- In this thesis different aspects of gel free procedures for mass spectrometry based proteome analysis are addressed, with emphasis on labelling techniques. It is based on three original articles where the effects imposed onto peptides upon modification are investigated, and the development of a new type of labelling strategy is described. Chemical modification has been of great importance for the peptide based proteomics approach, as they provide a way to impose desired properties onto the peptides. One possibility is to improve ionisation efficiency by introducing an easily protonated entity. By strategic placement of such a label, favoured ionisation of certain fragments upon MS/MS analysis can work to simplify the fragmentation spectra,... (More)
- In this thesis different aspects of gel free procedures for mass spectrometry based proteome analysis are addressed, with emphasis on labelling techniques. It is based on three original articles where the effects imposed onto peptides upon modification are investigated, and the development of a new type of labelling strategy is described. Chemical modification has been of great importance for the peptide based proteomics approach, as they provide a way to impose desired properties onto the peptides. One possibility is to improve ionisation efficiency by introducing an easily protonated entity. By strategic placement of such a label, favoured ionisation of certain fragments upon MS/MS analysis can work to simplify the fragmentation spectra, and consequently facilitate their interpretation. In this work, such a charge-directing fragmentation label and its effects using a new type of mass spectrometer was investigated. One of the main applications for the various labelling techniques however, has been the ability to produce isotopic counterparts of peptides, which enables quantification of the sample constituents. Several procedures to accomplish this, using a wide assortment of labels have, and are being developed. The Parent Ion Quantitation Scanning (PIQS) method described in this work, was developed aiming at more efficient global analysis of protein expression levels. The method was constructed to only sample peptides from differentially expressed proteins, thereby alleviating the large amounts of data usually generated, and only targeting relevant information, when aiming for analysis of differences in protein abundance. (Less)
- Abstract (Swedish)
- Popular Abstract in Swedish
I den här avhandlingen behandlas olika aspekter av gelfria metoder för proteomanalys med masspektrometri, med tonvikt på inmärkningsmetoder. Arbetet är baserat på tre artiklar, där olika effekter vid inmärkning på peptider utreds, samt framtagningen av en ny typ av inmärkningsstrategi beskrivs. Kemisk inmärkning har haft stor betydelse för peptidbaserad proteomik eftersom detta tillvägagångssätt möjliggör att olika eftersökta egenskaper kan tillföras på peptiderna. Ett exempel är möjligheten att förbättra joniseringsförmågan av peptider genom att förse dem med en modifiering som lätt kan protoneras. Genom strategisk placering av en sådan modifiering kan joniseringen av olika fragment vid MS/MS... (More) - Popular Abstract in Swedish
I den här avhandlingen behandlas olika aspekter av gelfria metoder för proteomanalys med masspektrometri, med tonvikt på inmärkningsmetoder. Arbetet är baserat på tre artiklar, där olika effekter vid inmärkning på peptider utreds, samt framtagningen av en ny typ av inmärkningsstrategi beskrivs. Kemisk inmärkning har haft stor betydelse för peptidbaserad proteomik eftersom detta tillvägagångssätt möjliggör att olika eftersökta egenskaper kan tillföras på peptiderna. Ett exempel är möjligheten att förbättra joniseringsförmågan av peptider genom att förse dem med en modifiering som lätt kan protoneras. Genom strategisk placering av en sådan modifiering kan joniseringen av olika fragment vid MS/MS analys påverkas. De fragment som bär på inmärkningen är fördelaktiga för jonisering, och därmed erhålls förenklade fragmenteringsspektra som underlättar tolkningen av dessa vid sekvensering. I den här avhandlingen har effekterna av en sort av denna inmärkning utvärderats med en ny typ av masspektrometer. En av de viktigaste tillämpningarna med kemisk inmärkning är dock möjligheten att generera motsvarande isotopa peptider, vilket möjliggör kvantifiering av komponenterna i ett prov. Flera olika metoder för att åstadkomma detta med olika typer av inmärkningar, har, och är under utveckling. Metoden ?Parent Ion Quantitation Scanning? (PIQS) vars framtagning beskrivs i denna avhandling, utvecklades med målsättningen att åstadkomma en mer effektiv, översiktlig analys av mängden uttryckta proteiner vid jämförelse mellan prov. Metoden konstruerades för att endast välja ut proteiner som skiljer sig i proteinmängd mellan två prov för analys. Med detta tillvägagångssätt minskas den enorma mängd data som ofta genereras, och endast relevant information filtreras ut. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/599123
- author
- Wåhlander, Åsa LU
- supervisor
-
- Peter James LU
- opponent
-
- Professor Doctor Vandekerckhove, Joël, University of Ghent, Belgien
- organization
- publishing date
- 2007
- type
- Thesis
- publication status
- published
- subject
- keywords
- Biotechnology, Bioteknik, Biokemisk teknik, Biochemical technology, 2D-HPLC, isotopes, labelling, proteomics, mass spectrometry
- pages
- 103 pages
- publisher
- Department of Immunotechnology, Lund University
- defense location
- Segerfalksalen, BMC, Sölvegatan 17, Lund
- defense date
- 2007-11-09 10:15:00
- ISBN
- 978-91-628-7328-8
- language
- English
- LU publication?
- yes
- id
- cd313c10-0f0d-4a9d-9514-c594ce6d2985 (old id 599123)
- date added to LUP
- 2016-04-04 10:01:36
- date last changed
- 2018-11-21 20:56:16
@phdthesis{cd313c10-0f0d-4a9d-9514-c594ce6d2985,
abstract = {{In this thesis different aspects of gel free procedures for mass spectrometry based proteome analysis are addressed, with emphasis on labelling techniques. It is based on three original articles where the effects imposed onto peptides upon modification are investigated, and the development of a new type of labelling strategy is described. Chemical modification has been of great importance for the peptide based proteomics approach, as they provide a way to impose desired properties onto the peptides. One possibility is to improve ionisation efficiency by introducing an easily protonated entity. By strategic placement of such a label, favoured ionisation of certain fragments upon MS/MS analysis can work to simplify the fragmentation spectra, and consequently facilitate their interpretation. In this work, such a charge-directing fragmentation label and its effects using a new type of mass spectrometer was investigated. One of the main applications for the various labelling techniques however, has been the ability to produce isotopic counterparts of peptides, which enables quantification of the sample constituents. Several procedures to accomplish this, using a wide assortment of labels have, and are being developed. The Parent Ion Quantitation Scanning (PIQS) method described in this work, was developed aiming at more efficient global analysis of protein expression levels. The method was constructed to only sample peptides from differentially expressed proteins, thereby alleviating the large amounts of data usually generated, and only targeting relevant information, when aiming for analysis of differences in protein abundance.}},
author = {{Wåhlander, Åsa}},
isbn = {{978-91-628-7328-8}},
keywords = {{Biotechnology; Bioteknik; Biokemisk teknik; Biochemical technology; 2D-HPLC; isotopes; labelling; proteomics; mass spectrometry}},
language = {{eng}},
publisher = {{Department of Immunotechnology, Lund University}},
school = {{Lund University}},
title = {{Chemical labelling methods for gel free based proteomics}},
year = {{2007}},
}