Purification and partial characterization of a novel phosphodiesterase from the venom of Trimeresurus stejnegeri: Inhibition of platelet aggregation

The phosphodiesterases (PDEs) are a superfamily of enzymes that have multiple roles in extracellular nucleotide metabolism and in the regulation of nucleotide-based intercellular signaling. Here we describe for the first time the isolation and partial characterization of a novel phosphodiesterase from Trimeresurus stejnegeri venom, named TS-PDE, using ion exchange and gel filtration chromatography. The purified TS-PDE is shown to be homogeneous as judged by SDS–PAGE and capillary isoelectric focusing. TS-PDE is a glycoprotein which contains 2.48% carbohydrate. Unlike other PDEs which are usually single polypeptide chain proteins with isoelectric points between 7.5 and 10.5, TS-PDE is a disulfide-linked heterodimer with an isoelectric point of 5.1 and a molecular mass of 100 kDa. The N-terminal amino acids of two chains are valine and serine, respectively. Furthermore, among all identified PDEs, only TS-PDE contains both of endogenous Cu2+ and Zn2+ which are essential for its phosphodiesterase activity. The purified TS-PDE exhibits broad phosphodiesterase substrate range with the order of specificity: nicotinamide guanine dinucleotide > ATP > nicotinamide adenine dinucleotide > ADP. The purified TS-PDE shows an exonuclease activity and no contamination with either alkaline phosphatase or 5′-nucleotidase activity. TS-PDE strongly inhibits ADP-induced platelet aggregation in human platelet-rich plasma by hydrolyzing ADP. Altogether, these results indicate that the novel TS-PDE is a unique phosphodiesterase with different structure from the known PDEs.

► A novel phosphodiesterase (TS-PDE) has been purified from Trimeresurus stejnegeri venom.
► Unlike other phosphodiesterases, TS-PDE is a disulfide-linked heterodimer with an isoelectric point of 5.1.
► Among all identified PDEs, only TS-PDE contains endogenous Cu2+ which is essential for its phosphodiesterase activity.
► TS-PDE exhibits broad phosphodiesterase substrate range with the order of specificity: NGD > ATP > NAD > ADP.
► TS-PDE shows an exonuclease activity and inhibits ADP-induced platelet aggregation by hydrolyzing ADP.