Molecular cloning of two toxic phospholipases A2 from the crown-of-thorns starfish Acanthaster planci venom

The full-length cDNAs encoding two toxic phospholipases A2 (AP-PLA2-I and -II) from the crown-of-thorns starfish Acanthaster planci venom were individually cloned by RT-PCR, 3′RACE and 5′RACE. In common with both AP-PLA2s, the precursor protein is composed of a signal peptide, a propeptide and a mature protein (136 and 135 residues for AP-PLA2-I and -II, respectively). The four motifs (Ca2+-binding loop, Ca2+-binding site, active site and catalytic network) characteristic of groups I and II PLA2s are well conserved in both AP-PLA2s. In addition to this, the presence of the elapid and pancreatic loops and the involvement of a propeptide in the precursors suggested that AP-PLA2s are highly analogous to the group IB PLA2s. However, when compared to the amino acid sequence of bovine pancreatic PLA2, the representative group IB PLA2, AP-PLA2s require some amino acid insertions and deletions in the region 76–100, as previously observed for the starfish Asterina pectinifera PLA2s. Furthermore, the phylogenetic tree made clearly demonstrated that AP-PLA2s and A. pectinifera PLA2s are distinguishable from the group IB PLA2s as well as other PLA2s, being classified into a new group.