Isolation and biochemical characterization of rubelase, a non-hemorrhagic elastase from Crotalus ruber ruber (Red Rattlesnake) venom

A novel non-hemorrhagic basic metalloprotease, rubelase, was isolated from the venom of Crotalus ruber ruber. Rubelase hydrolyzes succinyl-L-alanyl-L-alanyl-L-alanyl p-nitroanilide (STANA), a specific substrate for elastase, and the hydrolytic activity was inhibited by chelating agents. It also hydr...

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Bibliographic Details
Published in:Toxins 2011-07, Vol.3 (7), p.900-910
Main Authors: Komori, Yumiko, Sakai, Kaname, Masuda, Katsuyoshi, Nikai, And Toshiaki
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Amino Acids - analysis
Animals
Collagen - metabolism
Crotalid Venoms - genetics
Crotalid Venoms - isolation & purification
Crotalid Venoms - metabolism
Crotalus - metabolism
Crotalus ruber ruber toxin
cytotoxicity
elastase
Fibrinogen - metabolism
Hemorrhage
Human Umbilical Vein Endothelial Cells - drug effects
Human Umbilical Vein Endothelial Cells - metabolism
Humans
Hydrolysis
Metalloproteases - genetics
Metalloproteases - isolation & purification
Metalloproteases - metabolism
Molecular Sequence Data
Molecular Weight
Oligopeptides - metabolism
Pancreatic Elastase - genetics
Pancreatic Elastase - metabolism
Protein Conformation
Sequence Analysis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Venoms
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Description
Summary:A novel non-hemorrhagic basic metalloprotease, rubelase, was isolated from the venom of Crotalus ruber ruber. Rubelase hydrolyzes succinyl-L-alanyl-L-alanyl-L-alanyl p-nitroanilide (STANA), a specific substrate for elastase, and the hydrolytic activity was inhibited by chelating agents. It also hydrolyzes collagen and fibrinogen. However, hemorrhagic activity was not observed. By ESI/Q-TOF and MALDI/TOF mass spectrometry combined with Edman sequencing procedure, the molecular mass of rubelase was determined to be 23,266 Da. Although its primary structure was similar to rubelysin (HT-2), a hemorrhagic metalloprotease isolated from the same snake venom, the circumstances surrounding putative zinc binding domain HEXXHXXGXXH were found to be different when the three-dimensional computer models of both metalloproteases were compared. The cytotoxic effects of rubelase and rubelysin on cultured endothelial and smooth muscle cells were also different, indicating that the substitution of several amino acid residues causes the changes of active-site conformation and cell preference.
ISSN:2072-6651
2072-6651
DOI:10.3390/toxins3070900