Structure of bovine pancreatic phospholipase A2 at 1.7 Å resolution

Abstract

The crystal structure of bovine pancreatic phospholipase A2 has been refined to 1.7 Å resolution. The starting model for this refinement was the previously published structure at a resolution of 2.4 Å (Dijkstra et al., 1978). This model was adjusted to the multiple isomorphous replacement map with Diamond's real space refinement program (Diamond, 1971,1974) and subsequently refined using Agarwal's least-squares method (Agarwal, 1978). The final crystallographic R-factor is 17.1% and the estimated root-mean-square error in the positional parameters is 0.12 Å. The refined model allowed a detailed survey of the hydrogen-bonding pattern in the molecule. The essential calcium ion is located in the active site and is stabilized by one carboxyl group as well as by a peptide loop with many residues unvaried in all known phospholipase A2 sequences. Five of the oxygen ligands octahedrally surround the ion. The sixth octahedral position is shared between one of the carboxylate oxygens of Asp49 and a water molecule. The entrance to the active site is surrounded by residues involved in the binding of micelle substrates. The N-terminal region plays an important role here. Its α-NH + 3 group is buried and interacts with Gln4, the carbonyl oxygen of Asn71 and a fully enclosed water molecule, which provides a link between the N terminus and several active site residues. A total of 106 water molecules was located in the final structure, most of them in a two-layer shell around the protein molecule. The mobility in the structure was derived from the individual atomic temperature factors. Minimum mobility is found for the main chain atoms in the central part of the two long α-helices. The active site is rather rigid.