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

Abstract

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.

Modulation of the 5-Lipoxygenase Pathway by Chalcogen-Containing Inhibitors of Leukotriene A4 Hydrolase

“…Similar inhibition parameters with the three compounds with LTA 4 H may suggest that the hydrophobic interactions between the benzylphenyl moiety of an inhibitor and hydrophobic residues in LTA 4 H decide the binding efficiency. This is consistent with the structures of LTA 4 H co-crystallized with other benzylphenyl-containing inhibitors, 4-MDM [38] and 4-OMe-ARM1 [24]. Although 4-OMe-ARM1 shares a similar binding mode to ARM1, TTSe and TTO, the 4-MDM compound has the hydrogen-hydrogen interaction only between O of the methoxy group and Q136 in the middle of the substrate channel (Figure S8).…”

“…Until today, there have no comparative studies been performed to assess the efficacy of 4-MDM, ARM1-type inhibitors and resveratrol-type compounds in different disease models. It has been shown that LTA 4 H helps to prevent the accumulation of pro-inflammatory PGP in lungs [14,15,38]. Even though LTA 4 H-deficient mice lack both activities, it is not clear if higher levels of PGP in the absence of LTA 4 H are due to the lack of peptidase activity of LTA 4 H or compensating effects of the immune system [23].…”

Characterization of Aspergillus fumigatus secretome during sublethal infection of Galleria mellonella larvae

“…ITIH4 is linked to cell proliferation, as well as migration during the acute-phase inflammatory response, and serves a role in inflammatory and infectious responses, particularly in bacterial bloodstream infection [37]. Leukotriene A-4 hydrolase-like protein is an important inflammatory modulator, both promoting production of the pro-inflammatory mediator leukotriene B 4 but also degrading the neutrophil chemoattractant Pro-Gly-Pro (PGP) which could reduce inflammation [38,39]. The production of antimicrobial peptides such as cecropin-d-like peptide and gloverin-like protein-and fungal-specific recognition such as beta-1,3-glucan-binding protein-like isoform X2 was increased, indicating activation of an immune response of A. fumigatus proliferation.…”

Proteomic Analysis of Mesenchymal Stem Cells and Monocyte Co-Cultures Exposed to a Bioactive Silica-Based Sol–Gel Coating

“…The enzyme α-enolase (ENOA) stimulates the pro-inflammatory cytokine production (TNF-α, IL-1α/β, IFN-γ) and facilitates fibrinolysis by acting as a plasminogen receptor and degrading ECM . Other downregulated proteins were inhibitors of nuclear factor kappa-B kinase-interacting protein (IKIP), which is a key regulator of the nuclear factor kappa B (NF-kB) cascade, and leukotriene A 4 hydrolase (LKHA4), an inductor of neutrophilic infiltration in many inflammatory diseases . In addition, three cathepsins (CATB, CATZ, CATS), which have significant roles in the immune responses and are related to the NF-kB pathway, were also significantly affected.…”

“… 41 Other downregulated proteins were inhibitors of nuclear factor kappa-B kinase-interacting protein (IKIP), which is a key regulator of the nuclear factor kappa B (NF-kB) cascade, 42 and leukotriene A 4 hydrolase (LKHA4), an inductor of neutrophilic infiltration in many inflammatory diseases. 43 In addition, three cathepsins (CATB, CATZ, CATS), which have significant roles in the immune responses and are related to the NF-kB pathway, 44 were also significantly affected. Silica-based materials are known for inhibiting M1 polarization (pro-inflammatory) by reducing the expression of specific cell surface markers and pro-inflammatory cytokine production.…”