Iščitavanje reverzne translacije sekvenciranjem peptida de novo tehnikama tandemne spektrometrije masa

doctoral thesis

• Title (english): Scanning of reverse translation via de novo peptide sequencig by tandem mass spectrometry
• Author: Ivana Dodig
• Mentor: Mario Cindrić (mentor)
• Committee member: Ita Gruić Sovulj (predsjednik povjerenstva)
• Committee member: Mario Cindrić (član povjerenstva)
• Committee member: Predrag Novak (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2016-07-22, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry
• Universal decimal classification (UDC): 54 - Chemistry. Crystallography. Mineralogy

Abstract

Sekvenciranje peptida i proteina de novo korišteno je kao alat za reverzno
iščitavanje translacije. Pri tome je nedvojbeno sekvenciranje de novo omoguceno selektivnim
obilježavanjem N-kraja peptida 5-formil-1,3-benzen-di-sulfonicnom kiselinom prije analize
tandemnom spektrometrijom masa.
Opisana kemijska modifikacija usmjerava fragmentaciju peptidnih iona ka disocijaciji
peptidne veze prilikom kolizijom inducirane disocijacije u spektrometru masa. Nastali
fragmentni ioni b-serije... More detektiraju se u negativnom načinu rada spektrometra masa, a
fragmentni ioni y-serije detektiraju se u pozitivnom načinu spektrometra masa. Stoga je, za
isti peptid, aminokiselinski slijed iščitan iz spektra snimljenog u pozitivnom načinu rada
spektrometra masa moguće dodatno provjeriti iščitavanjem iz spektra snimljenog pri
negativnom načinu rada spektrometra masa (tzv. nedvojbeno sekvenciranje peptida de novo).
Jedna od primjena ovakvog načina sekvenciranja de novo je i identifikacija proteina, koja se
provodi pretragom, iz spektra iščitane sekvence peptida, u bazi podataka, odnosno
iščitavanjem reverzne translacije.
Disertacija prikazuje primjenjivost, pouzdanost i točnost kao i komparativnu prednost
novorazvijene de novo tehnike nad klasičnim tehnikama identifikacije proteina. Less

Abstract (english)

De novo peptide and protein sequencing was used as a tool for reverse translation
scanning. Unambiguous de novo sequencing is achived by selective derivatization of peptide
N-terminus by 5-formyl-1,3-benzendisulfonic acid before tandem mass spectrometry analysis.
Described chemical modification of peptide directs fragmentation pathway to the peptide
bound dissociation during collision-induced dissociation in the mass spectrometer. The
resulting b-series of fragmentation ions is... More detected in the negative ion mode of the mass
spectrometer, and y-series of fragmentation ions is detected in the positive ione mode of the
mass spectrometer. Therefore, for the same peptide, amino acid sequence read out from the
spectrum recorded in the positive ione mode can be additionally verified by reading out from
the spectrum obtained in the negative ion mode of mass spectrometer (the so-called
unambiguous de novo sequencing).
One application of the described de novo sequencing technique is protein identification, which
is performed by searching for previously spectral obtained peptide sequence, in the protein
database ie. by scanning of reverse translation.
This thesis presents applicability, reliability and accuracy as well as the comparative
advantage of the newly developed de novo sequencing technique over the conventional
protein identification techniques. Less

• Keywords: sekvenciranje de novo; identifikacija proteina; kemijski akivirana fragmentacija; tandemna spektrometrija masa; 5-formil-1; 3-benzen-di-sulfoniča kiselina
• Keywords (english): chemically activated fragmentation; de novo sequeincing; protein identification; tandem mass spectrometry; 5-formyl-1; 3-benzendisulfonic acid
• Language: croatian
• URN:NBN: urn:nbn:hr:217:611174
• 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: 191 str. ; 30 cm
• File origin: Born digital
• Access conditions: Open access
• Created on: 2017-03-09 10:23:08