A peptidomimetic fluorescent probe to detect the trypsin β2 subunit of the human 20S proteasome

Magdalena Wysocka , Anita Romanowska , Natalia Gruba , Michalina Michalska , Artur Giełdoń , Adam Lesner


This work describes the chemical synthesis, combinatorial selection, and enzymatic evaluation of peptidomimetic fluorescent substrates specific for the trypsin-like (β2) subunit of the 20S human proteasome. After deconvolution of a library comprising nearly 6000 compounds composed of peg substituted diaminopropionic acid DAPEG building blocks, the sequence ABZ–Dap(O2(Cbz))–Dap(GO1)–Dap(O2(Cbz))–Arg–ANB–NH2, where ABZ is 2-aminobenzoic acid, and ANB- 5 amino 2- nitro benzoic acid was selected. Its cleavage followed sigmoidal kinetics, characteristic for allosteric enzymes, with Km = 3.22 ± 0.02 μM, kcat = 245 s−1, and kcat/Km = 7.61 × 107 M−1 s−1. This process was practically halted when a selective inhibitor of the β2 subunit of the 20S human proteasome was supplemented to the reaction system. Titration of the substrate resulting in decreased amounts of proteasome 20S produced a linear signal up to 10−11 M. Using this substrate, we detected human proteasome 20S in human urine samples taken from the bladders of cancer patients. This observation could be useful for the noninvasive diagnosis of this severe disease.
Author Magdalena Wysocka (FCh/DBCh/LMCh)
Magdalena Wysocka,,
- Laboratory of Medical Chemistry
, Anita Romanowska (FCh/DBCh/LMCh)
Anita Romanowska,,
- Laboratory of Medical Chemistry
, Natalia Gruba (FCh/DET/LBAN)
Natalia Gruba,,
- Laboratory of Biochemical Analytics and Nanodiagnostics
, Michalina Michalska (FCh/DBCh/LMCh)
Michalina Michalska,,
- Laboratory of Medical Chemistry
, Artur Giełdoń (FCh/DTCh/LSP)
Artur Giełdoń,,
- Laboratory of Simulation of Polymers
, Adam Lesner (FCh/DET/LBAN)
Adam Lesner,,
- Laboratory of Biochemical Analytics and Nanodiagnostics
Journal seriesInternational Journal of Molecular Sciences, ISSN 1422-0067, (N/A 140 pkt)
Issue year2020
Publication size in sheets0.75
Article number2396
Keywords in Englishpeptidomimetics, libraries, fluorogenic substrates, proteasome, bladder cancer
ASJC Classification2700 General Medicine; 1312 Molecular Biology; 1503 Catalysis; 1604 Inorganic Chemistry; 1605 Organic Chemistry; 1606 Physical and Theoretical Chemistry; 1607 Spectroscopy; 1706 Computer Science Applications
Languageen angielski
LicenseRepository; published final; Uznanie Autorstwa (CC-BY); with publication
Wysocka_Magdalena_A_peptidomimetic_fluorescent_probe_2020.pdf Wysocka_Magdalena_A_peptidomimetic_fluorescent_probe_2020.pdf 6,29 MB
Score (nominal)140
Score sourcejournalList
ScoreMinisterial score = 140.0, 07-08-2020, ArticleFromJournal
Publication indicators WoS Citations = 0.000; Scopus SNIP (Source Normalised Impact per Paper): 2018 = 1.224; WoS Impact Factor: 2018 = 4.183 (2) - 2018=4.331 (5)
Citation count*
Additional fields
LicencjaUtwór jest udostępniany na licencji Creative Commons Uznanie autorstwa 4.0 Międzynarodowe (CC BY 4.0) https://creativecommons.org/licenses/by/4.0/d
Share Share

Get link to the record

* presented citation count is obtained through Internet information analysis and it is close to the number calculated by the Publish or Perish system.
Are you sure?