Separation of Crotalus atrox (Western diamondback rattlesnake) α-proteinase from serine proteinase and hemorrhagic factor activities

✍ Scribed by Marian Kruzel; Lawrence F. Kress

• Book ID: 102988329
• Publisher: Elsevier Science
• Year: 1985
• Tongue: English
• Weight: 646 KB
• Volume: 151
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(85)90207-6

✦ Synopsis

A method for obtaining Crotalus atrox a-proteinase (EC 3.4.24. I) in a pure form has been developed. Fractionation of the crude venom on DEAE-Sepharose, followed by gel filtration on Bio-Gel P-I50 and chromatography on CM-Sepharose, yielded an a-proteinase preparation which showed a single band on disc and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and had an activity on casein approximately twice that previously reported. The enzyme is a nonglycosylated single-chain polypeptide with a molecular weight of 26,738 and a pl of 8.15. Proteolytic activityoncasein,a,-antichymotrypsin,andCi-inhibitorwasabolishedbytreatmentofn-proteinase with I mM EDTA, but full activity was retained in the presence of I mM phenylmethylsulfonyl fluoride. Caseinolytic activity was increased by 33 and 55% in the presence of 10 mM M$+ and Ca*+, respectively. Pure cY-proteinase is devoid of esterolytic activity on H-D-Pro-Phe-Arg-p nitroanilide (S-2302) benzoyl+-arginine ethyl ester, and benzoyl+tyrosine ethyl ester. The final preparation has no hemorrhagic factor activity. o 1985 Academic press, h.