PhD Scientific Days 2023
Budapest, 22-23 June 2023
Pharmaceutical Sciences - Posters F
Introduction of a New Alkylating Agent for the Mass Spectrometry-Based Absolute Quantification of Proteins and Glycoproteins
Előadó neve
Dr. Virág, Dávid, PhD
Neptun code
GM8M7W
Előadó munkahelye
Semmelweis University Department of Pharmaceutics
Előadó telefonszáma
+36309402979
Előadó e-mail címe
virag.david@semmelweis.hu
Az előadás címe
Introduction of a New Alkylating Agent for the Mass Spectrometry-Based Absolute Quantification of Proteins and Glycoproteins
Szerző(k) neve és munkahelye
Dávid Virág1, Adina Borbély2, Gitta Schlosser2, Sára Berek1, Teodóra Borhegyi1, István Antal1, Borbála Dalmadi Kiss1, Krisztina Ludányi1
1 Department of Pharmaceutics, Semmelweis University, Hőgyes Endre utca 7., H-1092, Budapest, Hungary
2 MTA-ELTE Lendület Ion Mobility Mass Spectrometry Research Group, ELTE Eötvös Loránd University, Faculty of Science, Institute of Chemistry, Pázmány Péter sétány 1/A, H-1117, Budapest, Hungary
Bemutatás módja
Poszter
Szekció
Pharmaceutical Sciences - Posters F
Language of the presentation
Hungarian
Preferred session
Pharmaceutical Sciences
Összefoglaló szövege
Mass spectrometry (MS) has already become the technique of choice for analyzing proteins, however, absolute quantitative methods are typically expensive and labor-intensive. Consequently, a growing demand for simple yet reliable quantification approaches is witnessed.
Aim of the present work has been to develop a new quantitative proteomic strategy based on the differential alkylation of cysteine residues and evaluate its performance in the context of serum level determination and glycoform profiling.
Alpha-1-acid glycoprotein (AGP) was used as a model for the studies. Human serum and isolated protein samples were reduced and alkylated with iodacetic acid (IAA). Standard AGP was treated in a similar manner but changing the alkylating agent to potential alternatives such as iodopropionic acid (IPA) and iodobutyric acid (IBA). The samples and internal standards were than combined and digested enzymatically. The digests were cleaned up using solid phase extraction and acetonic precipitation. The samples were analyzed by means of liquid chromatography-tandem MS.
Among the alkylating agents tested IPA outperformed significantly IBA in terms alkylation efficiency. It was also shown that IAA and IPA labeled peptides had same extraction recoveries and ionization efficiencies. Since IPA introduces a 14 Da mass shift to the peptides, isotopic peak clusters of the analytes and internal standards are completely separated facilitating data analysis.
The results presented in this study support that cysteine-containing proteins labeled with IPA may serve as great internal standards enabling absolute quantitative analyses. In comparison with the available ones, the new strategy introduced seems to be simple, cost-effective and can be easily integrated in standard proteomic workflows. Therefore, it may be advantageously exploited in fields where reliable protein and glycoprotein quantitation is required such as biochemical studies, biomarker discovery and the characterization of protein biopharmaceuticals.
Project no. TKP2021-EGA-24 has been implemented with the support provided by the Ministry of Innovation and Technology of Hungary from the National Research, Development and Innovation Fund, financed under the TKP2021-EGA/TKP2021-NVA/TKP2021-NKTA funding scheme. DV was supported by the New National Excellence Program of the Ministry of Human Capacities (ÚNKP-22-4-II-SE-26).
University and Doctoral School
Semmelweis University, Doctoral School of Pharmaceutical Sciences
Supervisor
Dr. Krisztina Ludányi
Publication of my abstract
I do not give consent to the publication of my abstract on the website of the congress.
Kind
Szabad
Status
elfogadva
Accepted presentation method
poszter
Előadás fájl jóváhagyás
nem rendelkezett róla
Előadó
6056
Start
12:18
End
12:23
Authors (legacy)
Dávid Virág1, Adina Borbély2, Gitta Schlosser2, Sára Berek1, Teodóra Borhegyi1, István Antal1, Borbála Dalmadi Kiss1, Krisztina Ludányi1
1 Department of Pharmaceutics, Semmelweis University, Hőgyes Endre utca 7., H-1092, Budapest, Hungary
2 MTA-ELTE Lendület Ion Mobility Mass Spectrometry Research Group, ELTE Eötvös Loránd University, Faculty of Science, Institute of Chemistry, Pázmány Péter sétány 1/A, H-1117, Budapest, Hungary