Purification, characterization, and cDNA sequence of halysetin, a disintegrin-like/cysteine-rich protein from the venom of Agkistrodon halys Pallas.
By means of DEAE-Sepharose CL-6B column chromatography, gel filtration on Sephadex G-75 and Superose 12 FPLC, halysetin, an antiplatelet protein, was purified from the venom of Agkistrodon halys Pallas with molecular mass of 29 kDa on SDS-PAGE and 23,168 Da by mass spectrometry. The p1 was about 5.0. Halysetin was devoid of phospholipase A2, fibrino-(geno)lytic, esterase, hemorrhagenic activities. Halysetin dose-dependently inhibited the aggregation of human platelet, which was stimulated by collagen with IC50 of 420 nM, but not that stimulated by ADP. The N-and C-terminal sequences of halysetin were characterized. Its full-length cDNA was cloned by RT-PCR from the total RNA extracted from the snake venom gland. It encoded a protein of 212-amino-acid residues with disintegrin-like/cysteine-rich domains and was highly homologous with SYMPs (snake venom metalloprotease).
Liu
J W
JW
Institute of Biochemistry, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
Du
X Y
XY
Liu
P
P
Chen
X
X
Xu
J M
JM
Wu
X F
XF
Zhou
Y C
YC
D000244:Adenosine Diphosphate / Q000378:metabolism; D017836:Agkistrodon; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002847:Chromatography, Agarose; D002853:Chromatography, Liquid; D003094:Collagen / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry*; D018076:DNA, Complementary / Q000378:metabolism; D019483:Disintegrins / Q000737:chemistry*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004950:Esterases / Q000378:metabolism; D005337:Fibrin / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D010974:Platelet Aggregation; D012313:RNA / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012910:Snake Venoms / Q000737:chemistry; D014757:Viper Venoms / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*
D018076:DNA, Complementary; D019483:Disintegrins; D012910:Snake Venoms; D014757:Viper Venoms; C417253:halysetin; D000244:Adenosine Diphosphate; D012313:RNA; D005337:Fibrin; D005340:Fibrinogen; D003094:Collagen; D004950:Esterases; D010741:Phospholipases A; D054467:Phospholipases A2; D003545:Cysteine