Characterization of the isoforms of phospholipase A2 from honeybee venom

Abstract Phospholipase A 2 from the venom of the European honeybee ( Apis mellifera ) consists of three isoforms with approximate molecular masses of 16, 18, and 20 kDa, respectively, as deduced from SDS-PAGE. These variants, termed PLA-16, PLA-18, and PLA-20, were isolated by lectin affinity chromatography and preparative polyacrylamide gel electrophoresis. The amino acid sequences of the N -terminal peptide portions of all three isoforms, as assessed by automated Edman degradation, were identical with that expected for honeybee phospholipase A 2 . Sequencing data suggest that, while PLA-18 and PLA-20 carry oligosaccharide residues at asparagine-13, PLA-16 has escaped glycosylation during biosynthesis. Release of the carbohydrate from PLA-18 and PLA-20 with peptide: N -glycosidase F abolished the molecular mass differences between the three isoforms of phospholipase. Differences in sensitivity to α-mannosidase and monosaccharide composition of PLA-18 and PLA-20 further indicate that their electrophoretic separation is based on structural features of the N-glycosidically linked oligosaccharide. Noticeably, PLA-20 contains N -acetylgalactosamine, a sugar not having yet been described as a constituent of insect glycoproteins.