Unusual venom phospholipases A2 of two primitive tree vipers Trimeresurus puniceus and Trimeresurus borneensis.
To explore the venom diversity of Asian pit vipers, we investigated the structure and function of venom phospholipase A2 (PLA2) derived from two primitive tree vipers Trimeresurus puniceus and Trimeresurus borneensis. We purified six novel PLA2s from T. puniceus venom and another three from T. borneensis venom. All cDNAs encoding these PLA2s except one were cloned, and the molecular masses and N-terminal sequences of the purified enzymes closely matched those predicted from the cDNA. Three contain K49 and lack a disulfide bond at C61-C91, in contrast with the D49-containing PLA2s in both venom species. They are less thermally stable than other K49-PLA2s which contain seven disulfide bonds, as indicated by a decrease of 8.8 degrees C in the melting temperature measured by CD spectroscopy. The M110D mutation in one of the K49-PLA2s apparently reduced its edematous potency. A phylogenetic tree based on the amino-acid sequences of 17 K49-PLA2s from Asian pit viper venoms illustrates close relationships among the Trimeresurus species and intergeneric segregations. Basic D49-PLA2s with a unique Gly6 substitution were also purified from both venoms. They showed edema-inducing and anticoagulating activities. It is notable that acidic PLA2s from both venoms inhibited blood coagulation rather than platelet aggregation, and this inhibition was only partially dependent on enzyme activity. These results contribute to our understanding of the evolution of Trimeresurus pit vipers and the structure-function relationships between various subtypes of crotalid venom PLA2.
Wang
Ying-Ming
YM
Institute of Biological Chemistry, Academia Sinica and Institute of Biochemical Sciences, National Taiwan University, Taipei, Taiwan.
Peng
Hao-Fan
HF
Tsai
Inn-Ho
IH
D000595:Amino Acid Sequence; D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology; D001208:Asia; D001777:Blood Coagulation / Q000187:drug effects; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003435:Crotalid Venoms / Q000201:enzymology*; D004305:Dose-Response Relationship, Drug; D004487:Edema / Q000139:chemically induced; D004795:Enzyme Stability; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology*; D054467:Phospholipases A2; D010802:Phylogeny; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D017208:Rats, Wistar; D017421:Sequence Analysis; D013329:Structure-Activity Relationship; D017841:Trimeresurus
D000925:Anticoagulants; D003435:Crotalid Venoms; D010975:Platelet Aggregation Inhibitors; D010741:Phospholipases A; D054467:Phospholipases A2