Substrate-dependent modulation of the leukotriene A4 hydrolase aminopeptidase activity and effect in a murine model of acute lung inflammation

The aminopeptidase activity (AP) of the leukotriene A₄ hydrolase (LTA₄H) enzyme has emerged as a therapeutic target to modulate host immunity. Initial reports focused on the benefits of augmenting the LTA₄H AP activity and clearing its putative pro-inflammatory substrate Pro-Gly-Pro (PGP). However, recent reports have introduced substantial complexity disconnecting the LTA₄H modulator 4-methoxydiphenylmethane (4MDM) from PGP as follows: (1) 4MDM inhibits PGP hydrolysis and subsequently inhibition of LTA₄H AP activity, and (2) 4MDM activates the same enzyme target in the presence of alternative substrates. Differential modulation of LTA₄H by 4MDM was probed in a murine model of acute lung inflammation, which showed that 4MDM modulates the host neutrophilic response independent of clearing PGP. X-ray crystallography showed that 4MDM and PGP bind at the zinc binding pocket and no allosteric binding was observed. We then determined that 4MDM modulation is not dependent on the allosteric binding of the ligand, but on the N-terminal side chain of the peptide. In conclusion, our study revealed that a peptidase therapeutic target can interact with its substrate and ligand in complex biochemical mechanisms. This raises an important consideration when ligands are designed to explain some of the unpredictable outcomes observed in therapeutic discovery targeting LTA₄H.