Activation of bee venom phospholipase A2 by oleoyl imidazolide produces a thiol- and proteinase-resistant conformation.
Assay methods for bee venom phospholipase A2 are presented which respond to different aspects of enzymic behaviour and which allow basal activity, fatty acid activation and acyl-group activation to be distinguished. The stability of the enzyme to thiols and proteinases is dramatically increased by activation with the selective acylating agent, oleoyl imidazolide. These results support the model of activation by conformation change. Limited-fixation studies indicate that enzyme conformation is determined by interaction with the substrate. The oleoyl-enzyme is partially inactivated by trypsin, but its electrophoretic mobility is unchanged. This protective effect is highly selective and only one other component of the venom is protected against trypsin by oleoyl imidazolide. Combination of trypsin and thiol treatment produces a large fragment of the activated enzyme which could be used for structural studies of the activation site.
D001514:Bee Venoms; D004586:Electrophoresis; D010450:Endopeptidases / Q000494:pharmacology; D004789:Enzyme Activation; D007093:Imidazoles / Q000494:pharmacology*; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011487:Protein Conformation; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D001514:Bee Venoms; D007093:Imidazoles; D013438:Sulfhydryl Compounds; C018353:oleoylimidazolide; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D010450:Endopeptidases