Spektrometrija masa fragmenata peptidnih iona nastalih djelovanjem radikala

doctoral thesis

• Title (english): Mass spectrometry of peptide fragment ions induced by radicals
• Author: Anita Horvatić; MBZ: 316795
• Mentor: Mario Cindrić (mentor)
• Committee member: Vlasta Drevenkar (predsjednik povjerenstva)
• Committee member: Mario Cindrić (član povjerenstva)
• Committee member: Boris Mildner (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2014-06-18, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry
• Universal decimal classification (UDC): 54 - Chemistry. Crystallography. Mineralogy

Abstract

Određivanje primarne strukture proteina temeljni je zahtjev većine proteomskih analiza. Za određivanje slijeda aminokiselina se, uz metode Edmanove odgradnje, koristi tandemna spektrometrija masa temeljena na fragmentaciji peptidnih iona. Fragmentacija uvelike ovisi o sastavu i broju aminokiselina te posttranslacijskim modifikacijama. Radi poboljšanja ionizacije i fragmentacije se, osim kemijskog obilježavanja peptida, koriste i inicijatori fragmentacije koji desolvatiraju peptidne ione ili generiraju radikale u plinovitoj fazi spektrometra masa. Tijekom izrade doktorske disertacije ispitana je kontrolirana fragmentacija peptida u izvoru iona spektrometra masa MALDI-TOF/TOF (matricom pomognuta ionizacija uz desorpciju laserskim zračenjem-analizator vremena leta/ analizator vremena leta) djelovanjem inicijatora fragmentacije iz skupine persulfata, peroksida, sulfona te nitrila, dodanih u smjesu peptida i matrice. Predloženi su mehanizmi fragmentacije peptidnih iona inicirani radikalima, koji su dodatno eksperimentalno potvrđeni fragmentacijom kemijski obilježenih peptida. Spektri masa peptida nakon dodatka inicijatora fragmentacije u prvom stupnju MS-analize odgovaraju tandemnom spektru masa uz korištenje kolizijske ćelije, a ione produkte moguće je fragmentirati u drugom stupnju analize, čime je omogućena višestruka tandemna spektrometrija masa (tzv. pseudo MS3).

Abstract (english)

Protein primary structure elucidation is the key requirement in proteomics analyses. Except the Edman degradation, unambiguous amino acid sequence determination can be regularly performed by usage of tandem mass spectrometry, based on peptide ion fragmentation. Protein characteristics such as size, amino acid composition or post-translational modifications affect fragmentation. For ionization and fragmentation efficacy improvements, except chemical modification of peptides, fragmentation initiators can be used. Fragmentation initiators facilitate desolvation and generate gas phase radicals in the mass spectrometer. In this thesis, controlled in source peptide ion fragmentation induced by addition of fragmentation initiators was studied by means of matrix-assisted desorption/ionization mass spectrometry (MALDI-TOF/TOF MS). Fragmentation initiators used were classified as persulfates, peroxides, sulphones and nitriles. Radical mechanisms of peptide ion fragmentation mediated by fragmentation initiators were elucidated and confirmed by orthogonal experiment - chemically modified peptides fragmentation. Mass spectra obtained in the first stage mass spectrometry after fragmentation initiators addition are almost identical to conventional tandem mass spectra produced by collision induced dissociation. Product ions formed in ion source can be further fragmented in the second step of MS analysis. This concept enables multiple tandem mass spectrometry (pseudo MS3).

• Keywords: inicijatori fragmentacije; ISD; MALDI; spektrometrija masa; radikalski mehanizam fragmentacije
• Keywords (english): fragmentation initiators; ISD; MALDI; mass spectrometry; radical mediated fragmentation mechanism
• Language: croatian
• URN:NBN: urn:nbn:hr:217:178919
• Study programme: Title: Doctoral study in chemistry; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica znanosti, područje prirodnih znanosti, polje kemija (doktor/doktorica znanosti, područje prirodnih znanosti, polje kemija)
• Type of resource: Text
• Extent: 126 str. ; 30 cm
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-02-14 15:14:35