A digestive phospholipase A(2) in midguts of tobacco hornworms, Manduca sexta L.
We hypothesized that phospholipase A(2) (PLA(2)) is a common feature of insect digestive physiology. PLA(2) hydrolyzes polyunsaturated fatty acids (PUFAs) associated with the sn-2 position of phospholipids (PLs). We describe here a PLA(2) from midgut contents of the tobacco hornworm, Manduca sexta. Our results indicate that the enzyme is sensitive to pH (inactivated at low pH), protein concentration (up to 1.6&mgr;g/&mgr;l), substrate concentration (up to 1.4nmoles/reaction), temperature (up to 30 degrees C), and incubation time. We also found that PLA(2) activity is higher in fed than in starved larvae, and enzyme activity is associated with the midgut contents, rather than the midgut epithelium of fed larvae. All known secretory PLA(2)s, except for a PLA(2) in venom of the marine snail, Conus magus, require high calcium concentrations for catalysis, but the Manduca PLA(2) appears to be calcium-independent, and it exhibits increased PLA(2) activity in the presence of a calcium-chelator, EGTA. In addition, the partially purified Manduca PLA(2) is not inhibited by the phospholipid analog, oleyloxyethylphosphorylcholine. These findings suggest that the Manduca digestive PLA(2) may represent another novel form of PLA(2).
Stanley
D W.
DW
Insect Biochemistry/Physiology Laboratory, Department of Entomology, University of Nebraska, Lincoln, USA
Sarath
G
G
Rana
R L.
RL