AGKISTRODON ACUTUS HEMOCOAGULASE-B

摘要

The present invention provides agkistrodon acutus hemocoagulase-B. The agkistrodon acutus hemocoagulase-B is high-activity hemocoagulase separated from agkistrodon acutus venom. The hemocoagulase is a single-stranded glycoprotein comprising 236 amino acids, and has an amino acid sequence shown as SEQ ID No. 1. The strand comprises 6 pairs of disulfide bonds; the molecular weight of SDS-PAGE is approximately 35kD; the molecular weight after deglycosylation is 26116.7Da; and an isoelectric point (pI) is 6.0. A hemocoagulase molecule is modified by heterozygotic polysaccharides at an Asn 77, 100, 229 site. The hemocoagulase is serine proteinase. The present invention further provides a separation and purification method of the hemocoagulase. The method comprise: removing undissolved substances by using pretreatment; performing anion-exchange column chromatography for two times and Sephdex-G75 molecular sieve chromatography for one time; obtaining an active elution peak; performing dialysis and lyophilization, so as to obtain the high-purity agkistrodon acutus venom hemocoagulase. The activity of the agkistrodon acutus venom hemocoagulase is not less than 2000U/mg; the purity in the HPLC analysis can exceed 95%; and the yield is 0.25% to 0.30% calculated according to the weight of an agkistrodon acutus venom raw material.