Biochemical characteristics of immune-associated phospholipase A2 and its inhibition by an entomopathogenic bacterium, Xenorhabdus nematophila
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An entomopathogenic bacterium, Xenorhabdus nematophila, induces an immunosuppression of target insects by inhibiting phospholipase A2 (PLA2) activity. Recently, an immune-associated PLA2 gene was identified from the red flour beetle, Tribolium castaneum. This study cloned this PLA2 gene in a bacterial expression vector to produce a recombinant enzyme. The recombinant T. castaneum PLA2 (TcPLA2) exhibited its characteristic enzyme activity with substrate concentration, pH, and ambient temperature. Its biochemical characteristics matched to a secretory type of PLA2 (sPLA2) because its activity was inhibited by dithiothreitol (a reducing agent of disulfide bond) and bromophenacyl bromide (a specific sPLA2 inhibitor) but not by methylarachidonyl fluorophosphonate (a specific cytosolic type of PLA2). The X. nematophila culture broth contained PLA2 inhibitory factor(s), which was most abundant in the media obtained at a stationary bacterial growth phase. The PLA2 inhibitory factor(s) was heat-resistant and extracted in both aqueous and organic fractions. Effect of a PLA2-inhibitory fraction on the immunosuppression of T. castaneum was equally comparable with that resulted from inhibition of the TcPLA2 gene expression by RNA interference.
KeywordsX. nematophila phospholipase A2 T. castaneum immunosuppression RNA interference
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