Purification, sequencing, and phylogenetic analyses of novel Lys-49 phospholipases A(2) from the venoms of rattlesnakes and other pit vipers.
Basic phospholipase A(2) homologs with Lys49 substitution at the essential Ca(2+)-binding site are present in the venom of pit vipers under many genera. However, they have not been found in rattlesnake venoms before. We have now screened for this protein in the venom of rattlesnakes and other less studied pit vipers. By gel filtration chromatography and RP-HPLC, Lys49-phospholipase-like proteins were purified from the venoms of two rattlers, Crotalus atrox and Crotalus m. molossus, and five nonrattlers, Porthidium nummifer, Porthidium godmani, Bothriechis schlegelii, Trimeresurus puniceus, and Trimeresurus albolabris. Their N-terminal amino acid sequences were shown to be characteristic for this phospholipase subfamily. The purified basic proteins from rattlesnakes caused myonecrosis and edema in experimental animals. We have also cloned the cDNAs and solved the complete sequences of four novel Lys49-phospholipases from the venom glands of C. atrox, P. godmani, B. schlegelii, and Deinagkistrodon acutus (hundred-pace). Phylogenetic analyses based on the amino acid sequences of 28 Lys49-phospholipases separate the pitviper of the New World from those of the Old World, and the arboreal Asiatic species from the terrestrial Asiatic species. The implications of the phylogeny tree to the systematics of pit vipers, and structure-function relationship of the Lys49-phospholipases are discussed.
Tsai
I H
IH
Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan. bc201@gate.sinica.edu.tw
Chen
Y H
YH
Wang
Y M
YM
Tu
M C
MC
Tu
A T
AT
D019943:Amino Acid Substitution; D000818:Animals; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003435:Crotalid Venoms / Q000737:chemistry / Q000201:enzymology*; D017839:Crotalus; D004487:Edema / Q000139:chemically induced; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010741:Phospholipases A / Q000737:chemistry* / Q000235:genetics* / Q000633:toxicity; D010802:Phylogeny; D051381:Rats; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D020539:Sequence Analysis, Protein; D013329:Structure-Activity Relationship; D017819:Viperidae
D003435:Crotalid Venoms; D010741:Phospholipases A