Method and technique development in peptide and protein mass Spectrometry
2000 (English)Doctoral thesis, comprehensive summary (Other academic)
Mechanistic studies and applicability of several methods and techniques for studying peptides and proteins with mass spectrometry are presented. A model for the protein molecular ion formation in explosive matrix-assisted plasma desorption mass spectrometry is proposed, involving a contribution from the chemical energy release during conversion of the explosive, an enhanced protonation through collisions with products from the explosive decay, and electron scavenging by other products.
The design and construction of an electrospray ionization time-of-flight mass spectrometer with orthogonal extraction, providing a mass accuracy of 10 ppm and a resolving power of 7,500 are shown. This performance greatly facilitates the interpretation of peptide fragment spectra and increases the validity of substance identifications. It is demonstrated that the fragmentation techniques nozzle-skimmer dissociation and partial acidhydrolysis are partially complementary and that a combination of these techniques increases the degree of peptide sequence coverage, and solves sequence ambiguities, in many cases.
Electron capture dissociation has been implemented on a commercially available Fourier transform ion cyclotron resonance (FTICR) mass spectrometer, resulting in cleaner and more informative peptide fragment spectra compared to other fragmentation techniques. A mechanism for the dissociation observed during storage of large ion populations inside anexternal radio-frequency hexapole, is proposed, involving increased ion radial velocities combined with ion-gas collisions. Finally, a new method is presented where the ultrahigh resolving power of the FTICR spectrometer is used for accurate determinations of the isotopic state of single amide hydrogens within peptides after solution phase deuterium/hydrogen exchange. This approach allows studies of protein structure on the amino acid residue specific level.
Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2000. , 53 p.
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1104-232X ; 518
Materials science, electrospray ionization, peptide sequencing, Fourier transform ion cyclotron resonance, hydrogen/deuterium exchange, protein structure
Research subject Molecular Biotechnology
IdentifiersURN: urn:nbn:se:uu:diva-438ISBN: 91-554-4681-7OAI: oai:DiVA.org:uu-438DiVA: diva2:164863
2000-04-14, lecture room 4101, The Ångström laboratory, Uppsala, Uppsala, 10:15 (English)