Mechanistic insights into CrCEP1 : a dual-function cysteine protease with endo- and transpeptidase activity

Van Midden, Katarina Petra (author), Mantz, Melissa (author), Fonović, Marko (author), Gazvoda, Martin (author), Svete, Jurij (author), Huesgen, Pitter F. (author), Hoorn, Renier A. L. van der (author), Klemenčič, Marina (author)

Proteases, essential regulators of plant stress responses, remain enigmatic in their precise functional roles. By employing activity-based probes for real-time monitoring, this study aimed to delve into protease activities in Chlamydomonas reinhardtii exposed to oxidative stress induced by hydrogen peroxide. However, our work revealed that the activity-based probes strongly labelled three non-proteolytic proteins—PsbO, PsbP, and PsbQ—integral components of photosystem II's oxygen-evolving complex. Subsequent biochemical assays and mass spectrometry experiments revealed the involvement of CrCEP1, a previously uncharacterized papain-like cysteine protease, as the catalyst of this labelling reaction. Further experiments with recombinant CrCEP1 and PsbO proteins replicated the reaction in vitro. Our data unveiled that endopeptidase CrCEP1 also has transpeptidase activity, ligating probes and peptides to the N-termini of Psb proteins, thereby expanding the repertoire of its enzymatic activities. The hitherto unknown transpeptidase activity of CrCEP1, working in conjunction with its proteolytic activity, unveils putative complex and versatile roles for proteases in cellular processes during stress responses.

• Language: English
• Keywords: proteolysis, activity-based probes, papain-like cysteine protease, transpeptidation, green algae
• Work type: Article
• Typology: 1.01 - Original Scientific Article
• Organization: FKKT - Faculty of Chemistry and Chemical Technology
• Publication status: Published
• Publication version: Version of Record
• Year: 2024
• Number of pages: 15 str.
• Numbering: Vol. 271, pt. 1, art. 132505
• PID: 20.500.12556/RUL-158222
• UDC: 577.15
• ISSN on article: 0141-8130
• DOI: 10.1016/j.ijbiomac.2024.132505
• COBISS.SI-ID: 197196291
• Publication date in RUL: 30.05.2024

Record is a part of a journal

• Title: International journal of biological macromolecules
• Shortened title: Int. j. biol. macromol.
• Publisher: Elsevier
• ISSN: 0141-8130
• COBISS.SI-ID: 25637888

Licences

• License: CC BY-NC 4.0, Creative Commons Attribution-NonCommercial 4.0 International
• Link: http://creativecommons.org/licenses/by-nc/4.0/
• Description: A creative commons license that bans commercial use, but the users don’t have to license their derivative works on the same terms.

Secondary language

• Language: Slovenian
• Keywords: proteoliza, sonde na osnovi aktivnosti, papainu podobna cisteinska proteaza, transpeptidacija, zelene alge