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Pregled bibliografske jedinice broj: 828848

Benefits of selective peptide derivatization with sulfonating reagent at acidic pH for facile matrix-assisted laser desorption/ionization de novo sequencing


Butorac, Ana; Solak Mekić, Meliha; Hozić, Amela; Diminić, Janko; Gamberger, Dragan; Nišavić, Marija; Cindrić, Mario
Benefits of selective peptide derivatization with sulfonating reagent at acidic pH for facile matrix-assisted laser desorption/ionization de novo sequencing // RCM. Rapid communications in mass spectrometry, 30 (2016), 14; 1687-1694 doi:10.1002/rcm.7594 (međunarodna recenzija, članak, znanstveni)


Naslov
Benefits of selective peptide derivatization with sulfonating reagent at acidic pH for facile matrix-assisted laser desorption/ionization de novo sequencing

Autori
Butorac, Ana ; Solak Mekić, Meliha ; Hozić, Amela ; Diminić, Janko ; Gamberger, Dragan ; Nišavić, Marija ; Cindrić, Mario

Izvornik
RCM. Rapid communications in mass spectrometry (0951-4198) 30 (2016), 14; 1687-1694

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
De novo sequencing ; peptide derivatization ; MALDI-TOF/TOF

Sažetak
RATIONALE One of the most challenging tasks of proteomics is peptide de novo sequencing. 4- Sulfophenyl isothiocyanate (SPITC) peptide derivatization enables acquisition of high- quality tandem mass spectra (MS/MS) for de novo sequencing, but unwanted non-specific reactions and reduced mass spectra (MS) signal intensities still represent the obstacles in high-throughput de novo sequencing. METHODS We developed a SPITC peptide derivatization procedure under acidic conditions (pH ≤5). Derivatized peptides were analyzed by matrix- assisted laser desorption/ionization (MALDI-MS) in negative ion mode followed by MS/MS in positive ion mode. A de novo sequencing tool, named DUST, adjusted to SPITC chemistry, was designed for successful high-throughput peptide de novo sequencing. This high-throughput peptide de novo sequencing was tested on Fusarium delphinoides, an organism with an uncharacterized genome. RESULTS The SPITC derivatization procedure under acidic conditions produced a significantly improved MS dataset in comparison to commonly used derivatization under basic conditions. Signal intensities were 6 to 10 times greater and the over-sulfonation effect measured on lysine- containing peptides was significantly decreased. Furthermore, development of a novel DUST algorithm enabled automated de novo sequencing with the calculated accuracy of 70.6%. CONCLUSIONS The SPITC derivatization and de novo sequencing approach outlined here provides a reliable method for high-throughput peptide de novo sequencing. High-throughput peptide de novo sequencing enabled protein mutation identification and identification of proteins from organisms with non-sequenced genomes.

Izvorni jezik
Engleski

Znanstvena područja
Kemija



POVEZANOST RADA


Ustanove
Institut "Ruđer Bošković", Zagreb

Časopis indeksira:


  • Current Contents Connect (CCC)
  • Web of Science Core Collection (WoSCC)
    • Science Citation Index Expanded (SCI-EXP)
    • SCI-EXP, SSCI i/ili A&HCI
  • Scopus
  • MEDLINE


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