0602578 - MBÚ 2025 RIV US eng J - Článek v odborném periodiku
Kalaninová, Zuzana - Portašiková, Jasmína Mária - Jirečková, Barbora - Polák, Marek - Nováková, J. - Kavan, Daniel - Novák, Petr - Man, Petr
Postproline Cleaving Enzymes also Show Specificity to Reduced Cysteine.
Analytical Chemistry. Roč. 96, č. 48 (2024), s. 19084-19092. ISSN 0003-2700. E-ISSN 1520-6882
Grant CEP: GA TA ČR(CZ) TN02000132; GA ČR(CZ) GA22-27695S; GA MŠMT(CZ) EH22_008/0004597; GA MŠMT(CZ) EF18_046/0015974
Výzkumná infrastruktura: CIISB III - 90242
Institucionální podpora: RVO:61388971
Klíčová slova: exchange-mass-spectrometry * prolyl endoprotease * peptide libraries * digestion * proteomics * proteases * hsp70 * tool
Obor OECD: Biochemistry and molecular biology
Impakt faktor: 6.7, rok: 2023 ; AIS: 1.224, rok: 2023
Způsob publikování: Open access
Web výsledku:
https://pubs.acs.org/doi/10.1021/acs.analchem.4c04277DOI: https://doi.org/10.1021/acs.analchem.4c04277

In proteomics, postproline cleaving enzymes (PPCEs), such as Aspergillus niger prolyl endopeptidase (AnPEP) and neprosin, complement proteolytic tools because proline is a stop site for many proteases. But while aiming at using AnPEP in online proteolysis, we found that this enzyme also displayed specificity to reduced cysteine. By LC-MS/MS, we systematically analyzed AnPEP sources and conditions that could affect this cleavage preference. Postcysteine cleavage was blocked by cysteine modifications, including disulfide bond formation, oxidation, and alkylation. The last modification explains why this activity has remained undetected so far. In the same experimental paradigm, neprosin mimicked this cleavage specificity. Based on these findings, PPCEs cleavage preferences should be redefined from post-Pro/Ala to post-Pro/Ala/Cys. Moreover, this evidence demands reconsidering PPCEs applications, whether cleaving Cys-rich proteins or assessing Cys status in proteins, and calls for revisiting the proposed enzymatic mechanism of these proteases.
Trvalý link: https://hdl.handle.net/11104/0359814


Vědecká data: Zenodo, Zenodo