Primary structure of a hemorrhagic metalloproteinase, HT-2, isolated from the venom of Crotalus ruber ruber.
Crotalidae and Viperidae snake venoms contains several kinds of metalloproteinases which cause localized hemorrhage by direct action on blood vessel walls. We report here the entire amino acid sequence and the disulfide bridge locations of HT-2, one of the hemorrhagic toxins isolated from the venom of Crotalus ruber ruber (red rattlesnake). The non-reduced protein was first cleaved at methionine residues to provide a set of 8 fragments, which covered the entire sequence of HT-2. The disulfide bridge locations of HT-2 were also determined by using these primary fragments. The unambiguous sequence for the whole protein was then established by conventional methods using lysyl endopeptidase and thermolysin digests. HT-2 consisted of 202 amino acid residues with two disulfide bridges, which were assigned to Cys-117-Cys-197 and Cys-157-Cys-164. HT-2 had a typical zinc-chelating sequence His-Glu-X-X-His (residues 142-146) found in thermolysin, and its overall sequence showed, respectively, 50, 52, and 53% identities to those of HR2a, H2-proteinase, and the metalloproteinase domain of HR1B. However, the disulfide bridge locations of HT-2 were different from those in the other metalloproteinases. The primary structure of HT-2 was more closely related to that of Ht-d from Crotalus atrox recently determined (81% identity). From the structural comparison with five metalloproteinases so far elucidated, six conservative amino acid residues, which may possibly be related to the induction of the hemorrhagic activity, were suggested to be present in these toxins.
Takeya
H
H
Department of Molecular Biology, Graduate School of Medical Science, Fukuoka.
Onikura
A
A
Nikai
T
T
Sugihara
H
H
Iwanaga
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003435:Crotalid Venoms / Q000737:chemistry*; D003488:Cyanogen Bromide; D004220:Disulfides / Q000737:chemistry; D006868:Hydrolysis; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011487:Protein Conformation; D012911:Snakes
D003435:Crotalid Venoms; D004220:Disulfides; D008666:Metalloendopeptidases; C044577:atrolysin A; D003488:Cyanogen Bromide