Wydział Chemii

Remigiusz Bąchor

2022

Molecules

27

4107/1-4107/11

10.3390/molecules27134107

Naukowa

Angielski

Artykuł

Caspases, proteolytic enzymes belonging to the group of cysteine proteases, play a crucial
role in apoptosis. Understanding their activity and substrate specificity is extremely important.
Fluorescence-based approaches, including fluorogenic substrates, are generally used to confirm
cleavage preferences. Here we present a new method of substrate specificity and activity analysis
based on the application of fix-charge tagged peptides located on the resin. The proteolysis of peptide
bond on the resin, occurring even with low efficiency, results in the formation of N-terminal fragments
of model peptide containing ionization enhancers in the form of quaternary ammonium groups,
allowing for ultrasensitive and reliable analysis by LC-MS/MS. The possibility of application of the
proposed solution was tested through the analysis of substrate specificity and activity of caspase 3 or
7. The obtained results confirm the known substrate specificity of executioner caspases. Our solution
also allowed us to observe that caspases can hydrolyze peptides shorter than those presented to date
in the scientific literature.

caspases, substrate specificity, on-bead proteolysis, OBOC, fixed-charge tag, ESI-MS

CC-BY

http://dx.doi.org/10.3390/molecules27134107

http://www.mdpi.com/journal/metals