Crystal structure of a carbohydrate induced homodimer of phospholipase A₂ from Bungarus caeruleus at 2.1 Å resolution

Abstract

This is the first crystal structure of a carbohydrate induced dimer of phospholipase A₂ (PLA₂). This is an endogenous complex formed between two PLA₂ molecules and two mannoses. It was isolated from Krait venom (Bungarus caeruleus) and crystallized as such. The complete amino acid sequence of PLA₂ was determined using cDNA method. Three-dimensional structure of the complex has been solved with molecular replacement method and refined to a final R-factor of 0.192 for all the data in the resolution range 20.0-2.1 Å. The presence of mannose molecules in the protein crystals was confirmed using dinitrosalicylic acid test and the molecular weight of the dimer was verified with MALDI-TOF. As indicated by dynamic light scattering and analytical ultracentrifugation the dimer was also stable in solution. The good quality non-protein electron density at the interface of two PLA₂ molecules enabled us to model two mannoses. The mannoses are involved extensively in interactions with protein atoms of both PLA₂ molecules. Some of the critical amino acid residues such as Asp 49 and Tyr 31, which are part of the substrate-binding site, are found facing the interface and interacting with mannoses. The structure of the complex clearly shows that the dimerization is caused by mannoses and it results in the loss of enzymatic activity.