Kinase experiment:

The ability of PL553 to discriminate LTA4H from APN enzymatic activity is tested using the (l)-Ala-β-naphthylamide and PL553 peptide substrates at 40 μM with either APN from porcine kidney (0.33 mU/mL) or recombinant human LTA4H (0.6 μg/mL). The enzymatic reactions proceeds for 1 h at 37℃ in a final volume of 100 μL of 50 mM Tris-HCl (pH 7.4) or 50 mM Tris-HCl (pH 7.4) and 100 mM NaCl for APN or LTA4H, respectively[1].

PL553 is a specific and high-affinity fluorigenic substrate of Leukotriene A4 hydrolase, with a λmax of 210 nm and λem of 410 nm.

PL553 is a specific and high-affinity fluorigenic substrate of Leukotriene A4 hydrolase (LTA4H), with a maximum absorption (λmax) of 210 nm and maximum emission (λem) of 410 nm. PL553 is a better LTA4H substrate than (l)-Ala-β-naphthylamide, and resistant to cleavage by other aminopeptidases, but can be cleaved by FAAH. PL553 (40 μM) is used to evaluate the potencies of known inhibitors toward LTA4H[1].

[1]. Poras H, et al. A sensitive fluorigenic substrate for selective in vitro and in vivo assay of leukotriene A4 hydrolase activity. Anal Biochem. 2013 Oct 15;441(2):152-61.