Sk. Han et al., BACTERIAL EXPRESSION AND CHARACTERIZATION OF HUMAN PANCREATIC PHOSPHOLIPASE A(2), Biochimica et biophysica acta, L. Lipids and lipid metabolism, 1346(2), 1997, pp. 185-192
Mammalian pancreatic phospholipases A(2) (PLA(2)) have recently been i
mplicated in cell surface receptor-mediated inflammation. As a first s
tep toward understanding how human pancreatic PLA(2) (hp-PLA(2)) inter
acts with membranes and other biological targets including cell-surfac
e receptors, we constructed its bacterial expression vector which can
be used for the mutagenesis and protein over-expression. The expressio
n vector (pSH-hp) was constructed using a synthetic hp-PLA(2) gene who
se transcription is controlled by T7 promoter. hp-PLA(2) was expressed
as a mature protein in high concentration in Escherichia coli cells a
nd formed inclusion body. The solubilization of inclusion body protein
followed by the refolding and purification produced ca. 5 mg of pure
protein from one liter of growth medium. Kinetic studies of recombinan
t human, bovine and porcine pancreatic PLA(2)s using polymerized mixed
Liposomes and micelles as substrates showed that despite their highly
homologous structures these mammalian pancreatic PLA(2)s have distinc
t phospholipid head group specificity and different activity toward va
rious lipid substrates.