BIOCHEMICAL-CHARACTERIZATION OF DROSOPHILA-MELANOGASTER ACETYLCHOLINESTERASE EXPRESSED BY RECOMBINANT BACULOVIRUSES

Recombinant baculoviruses expressing full length and 3' truncated form s of c-DNA encoding the Drosophila melanogaster acetylcholinesterase ( AChE) were constructed. Biochemical analyses showed that full length r ecombinant protein was enzymatically active and anchored to the cell m embrane via a glycolipidic residue. DTT treatment dissociated the nati ve form into monomers migrating as did the corresponding form of AChE extracted from drosophila heads. Finally, DFP labelling demonstrated t hat the specific proteolytic cleavage leading to the formation of 55 a nd 16 kDa subunits occurred in Sf9 cells. In contrast with the full-le ngth enzyme, C-terminal-truncated forms were highly secreted, confirmi ng the prominent role of the C-terminal hydrophobic peptide for the ad dition of the glycolipidic residue. Accumulation of inactive precursor was observed when recombinant proteins were overproduced using an imp roved baculovirus, suggesting a saturation of insect cell machineries. (C) 1994 Academic Press, Inc.