A Midgut Digestive Phospholipase A2 in Larval Mosquitoes, Aedes albopictus and Culex quinquefasciatus.

Abstract

Phospholipase A₂ (PLA₂) is a secretory digestive enzyme that hydrolyzes ester bond at sn-2 position of dietary phospholipids, creating free fatty acid and lysophospholipid. The free fatty acids (arachidonic acid) are absorbed into midgut cells. Aedes albopictus and Culex quinquefasciatus digestive PLA₂ was characterized using a microplate PLA₂ assay. The enzyme showed substantial activities at 6 and 8 μg/μl of protein concentration with optimal activity at 20 and 25 μg/μl of substrate concentration in Aedes albopictus and Culex quinquefasciatus, respectively. PLA₂ activity from both mosquitoes increased in a linear function up to 1 hour of the reaction time. Both enzymes were sensitive to pH and temperature. PLA₂ showed higher enzyme activities in pH 8.0 and pH 9.0 from Aedes albopictus and Culex quinquefasciatus, respectively, at 40°C of incubation. The PLA₂ activity decreased in the presence of 5 mM (Aedes albopictus) and 0.5 mM (Culex quinquefasciatus) site specific PLA₂ inhibitor, oleyloxyethylphosphorylcholine. Based on the migration pattern of the partially purified PLA₂ on SDS-PAGE, the protein mass of PLA₂ is approximately 20-25 kDa for both mosquitoes. The information on PLA₂ properties derived from this study may facilitate in devising mosquitoes control strategies especially in the development of inhibitors targeting the enzyme active site.