학술논문
鹿茸菇子实体新纤溶酶的高效分离纯化 / Efficient isolation and purification of a novel fibrinolytic enzyme from fruiting bodies of Lyophyllum decastes
Document Type
Academic Journal
Author
Source
天然产物研究与开发 / Natural Product Research and Development. 35(11):1947-1938
Subject
Language
Chinese
ISSN
1001-6880
Abstract
采用层析分离法从食药兼用真菌鹿茸菇(Lyophyllum decastes)的子实体中纯化得到了一种新纤溶酶,采用Edman降解法测定了该酶N-端氨基酸序列.结果显示,鹿茸菇子实体经干燥、粉碎、浸提后获得的含纤溶酶的粗酶液,依次经过盐析、Octyl-Sepharose Fast Flow疏水相互作用层析、SP-Sepharose High Performance离子交换层析和Source 15PHE疏水相互作用层析分离后获得单一组分的纤溶酶,酶比活力达到了 4 105.78 U/mg,纯化倍数为206.09,酶活力回收率为30.91％.Native-PAGE电泳和SDS-PAGE电泳检测结果均表明,该纤溶酶纯度达到了电泳纯,相对分子量为30.9 kDa.Edman降解法测定纤溶酶的N-端氨基酸序列为:Gly-Ala-Val-Thr-Gln-Cys-Asn-Ala-Pro-Trp-Gly-Leu,经过NCBI数据库比对,该纤溶酶为一种新纤溶酶.本文的研究为纤溶酶的研发提供了新的思路和方法.
A novel fibrinolytic enzyme was isolated and purified from the fruiting bodies of Lyophyllum decastes.The N-termi-nal amino acid sequence of the enzyme was determined by Edman degradation method.The fruiting bodies of Lyophyllum de-castes was dried,crushed and extracted with phosphate solution to obtain crude enzyme solution.The crude enzyme solution was isolated and purified by using ammonium sulfate precipitation,Octyl-Sepharose Fast Flow hydrophobic chromatography,SP-Sepharose high performance ion chromatography and Source 15PHE hydrophobic chromatography to obtain a single compo-nent with fibrinolytic activity.The specific activity of the enzyme was 4 105.78 U/mg,the purification fold was 206.09,and the recovery rate of the activity was 30.91％.Native-PAGE and SDS-PAGE showed that the fibrinolytic enzyme reached elec-trophoretic purity and the molecular weight was 30.9 kDa.The N-terminal sequences of the fibrinolytic enzyme were deter-mined by the Edman degradation method,and the sequences was Gly-Ala-Val-Thr-Gln-Cys-Asn-Ala-Pro-Trp-Gly-Leu.By NCBI database comparison,it was found that the fibrinolytic enzyme was a novel fibrinolytic enzyme.The research provides a new idea and method for the research and development of fibrinolytic enzyme.
A novel fibrinolytic enzyme was isolated and purified from the fruiting bodies of Lyophyllum decastes.The N-termi-nal amino acid sequence of the enzyme was determined by Edman degradation method.The fruiting bodies of Lyophyllum de-castes was dried,crushed and extracted with phosphate solution to obtain crude enzyme solution.The crude enzyme solution was isolated and purified by using ammonium sulfate precipitation,Octyl-Sepharose Fast Flow hydrophobic chromatography,SP-Sepharose high performance ion chromatography and Source 15PHE hydrophobic chromatography to obtain a single compo-nent with fibrinolytic activity.The specific activity of the enzyme was 4 105.78 U/mg,the purification fold was 206.09,and the recovery rate of the activity was 30.91％.Native-PAGE and SDS-PAGE showed that the fibrinolytic enzyme reached elec-trophoretic purity and the molecular weight was 30.9 kDa.The N-terminal sequences of the fibrinolytic enzyme were deter-mined by the Edman degradation method,and the sequences was Gly-Ala-Val-Thr-Gln-Cys-Asn-Ala-Pro-Trp-Gly-Leu.By NCBI database comparison,it was found that the fibrinolytic enzyme was a novel fibrinolytic enzyme.The research provides a new idea and method for the research and development of fibrinolytic enzyme.