Activation and Inhibition of Leukotriene A4 Hydrolase Aminopeptidase Activity by Modulators B08-2a and B08-2b
: Leukotriene A4 hydrolase (LTA4H) is a bi-functional enzyme that promotes a pro-inflammatory epoxy hydrolase (EH) pathway and an anti-inflammatory aminopeptidase (AP) pathway. LTA4H EH activity catalyzes the hydrolysis of leukotriene A4 (LTA4) to the neutrophil chemoattractant leukotriene B4 (LTB4), while AP activity hydrolyzes the tripeptide proline-glycine-proline (PGP) to clear neutrophils in the lungs. Previously published studies focused on blocking EH activity to inhibit the LTB4 formation with small molecule inhibitors. Our research focuses on the activation of LTA4H AP activity in the presence of small molecule activators. Prior to kinetic analysis, a standard curve was obtained using para-nitroanilide (pNA) at escalating concentrations. A chromogenic substrate alanine-p-nitroanilide (Ala-pNA) was used to measure the AP activity of LTA4H. Using the 4-methoxydiphenylmethane (4MDM) as a positive control for activating the AP activity and bestatin as a negative control for inhibiting AP activity, the modulation of LTA4H AP activity was analyzed in the presence of compounds B08-2a and B08-2b through the determination of a 50% activation or inhibition (AC50/IC50) kinetic assay. In conclusion, our research involves the high-throughput assay for screening small molecules to modulate the AP activity of LTA4H. The discovery of small molecules that activate the AP activity of the LTA4H enzyme is a potential new therapeutic agent for lung inflammatory diseases, such as chronic obstructive pulmonary disease (COPD).
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