Purification, characterization and gene cloning of a novel phospholipase A2 from the venom of Agkistrodon blomhoffii ussurensis

A new phospholipase A2 with Gln at the site 49, abbreviated as Gln49-PLA2, has been purified from the venom of Agkistrodon blomhoffii ussurensis by using ion-exchange chromatography, gel filtration chromatography and reversed-phase HPLC, and behaves as a single-band on SDS-PAGE. Its molecular weight is 13881.85 ± 0.33 Da given by mass spectrometry and pI is about 8.56 given by isoelectric focusing. Gln49-PLA2 does not show phospholipase A2 and hemorrhagic activity, whereas shows weak toxic and apparent anticoagulant activity. Based on the N-terminal sequencing and peptide mass fingerprint analysis, Gln49-PLA2 cDNA has been cloned by means of RT-PCR. Gln49-PLA2 consists of 122 amino acid residues and has the structural features of class II of snake venom phospholipase A2.