A digestive phospholipase a2 in Midguts of worker honey bees, Apis mellifera : significance in lipid nutrition and health
Phospholipase A2 (PLA2) catalyzes hydrolysis of the fatty acid moiety from the sn-2 position of phospholipids. At least 16 groups of PLA2s are known in mammals. In insects, PLA2s occur in several tissues, where they act in a variety of biological functions, such as digestion, immune signaling, development and reproduction. However, other than the venom PLA2s, characterization of their physiological functions has not been reported in Hymenoptera, particularly in honey bees (Apis mellifera). Among many studies on honey bee nutrition and physiology, the enzymes acting in digesting dietary lipids remain understudied. This is an important lacuna because it bears directly on meeting essential dietary fatty acid requirements, which I addressed by characterizing a digestive PLA2 in midgut preparations of honey bee workers. The honey bee midgut PLA2 is sensitive to reaction conditions, including incubation time, substrate and enzyme concentrations, reaction temperature and pH. It is Ca2+- independent and susceptible to specific PLA2 inhibitors. Through a cryopreservation study of this PLA2 in honey bee midgut preparations, I confirmed that the honey bee digestive PLA2 preparations are physically stable, with high activity after storage at -80[degrees]C for at least three months. These findings open a new corridor of research into honey bee dietary lipid digestion and nutrition. I used classical molecular tools to identify genes encoding honey bee midgut PLA2s, to clone the genes from midgut tissue preparations and express one of the identified genes in competent E. coli cells. Although the enriched PLA2 from E. coli cultures was not biologically active, this work is a meaningful aspect of my training and education, on which I offer my thoughts in Chapter 4.
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