Design, synthesis, and characterization of chromogenic substrates of coagulation factor XIIIa

Abstract

A series of Glu(pNA)-containing peptides was designed to determine the activity of the transglutaminase factor XIIIa at 405 nm due to p-nitroaniline release. The most suitable substrate properties were found for peptides containing the Glu(pNA) residue in the second position from the N terminus. For the best substrate 12 (H-Tyr-Glu(pNA)-Val-Lys-Val-Ile-Gly-NH(2)), a k(cat)/K(m) value of 3531 s(-1)M(-1) was found. Although the k(cat)/K(m) values of the Glu(pNA) peptides are more than 100-fold reduced compared with the previously reported cleavage of natural glutamine-containing substrates such as α(2)-antiplasmin and β-casein, these chromogenic substrates can be useful tools for convenient determination of FXIII-A(2)* activity e.g., for in vitro inhibitor screening. As an example, peptide 12 was used to characterize the inhibition of FXIII-A(2)* by the well-known irreversible inhibitor iodoacetic acid.