Molecular cloning of two toxic phospholipases A 2 from the crown-of-thorns starfish Acanthaster planci venom

Abstract

The full-length cDNAs encoding two toxic phospholipases A 2 (AP-PLA 2-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-PLA 2s, the precursor protein is composed of a signal peptide, a propeptide and a mature protein (136 and 135 residues for AP-PLA 2-I and -II, respectively). The four motifs (Ca 2+-binding loop, Ca 2+-binding site, active site and catalytic network) characteristic of groups I and II PLA 2s are well conserved in both AP-PLA 2s. In addition to this, the presence of the elapid and pancreatic loops and the involvement of a propeptide in the precursors suggested that AP-PLA 2s are highly analogous to the group IB PLA 2s. However, when compared to the amino acid sequence of bovine pancreatic PLA 2, the representative group IB PLA 2, AP-PLA 2s require some amino acid insertions and deletions in the region 76–100, as previously observed for the starfish Asterina pectinifera PLA 2s. Furthermore, the phylogenetic tree made clearly demonstrated that AP-PLA 2s and A. pectinifera PLA 2s are distinguishable from the group IB PLA 2s as well as other PLA 2s, being classified into a new group.