Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Nereides protease, separating and purifying method and application thereof

A purification method and protease technology are applied in the fields of nerei protease, its separation, extraction and purification, and the separation, extraction and purification of plasmin, which can solve the problems of high price, short half-life, poor selectivity and the like.

Inactive Publication Date: 2004-06-02
洪敏
View PDF0 Cites 21 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The present invention provides a nereis protease, and at the same time provides a separation, extraction and purification method and application of the enzyme, which overcomes the disadvantages of conventional thrombolytic agents such as high price, poor selectivity, short half-life, and strong antigenicity, and can realize industrialization Production

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0112] Take 200g clamworm, wash and mince, add 4.5 times volume of pH7.0, 0.01mol / LNa 2 HPO 4 -NaH 2 PO 4 , the buffer solution was made into a homogenate; centrifuged at 9000 rpm for 30 minutes at 4°C, and the supernatant was obtained, which was the crude extract of N-V protease. Take 1000ml of N-V protease crude extract, put it in an ice bath and cool it to 0°C, add -20°C pre-cooled absolute ethanol under stirring conditions, so that the concentration of ethanol reaches 5%, keep it in an ice bath to cause precipitation ;Centrifuge at -10°C, 9000 rpm for 30 minutes, discard the precipitate; add -20°C pre-cooled absolute ethanol to the supernatant while stirring, the final concentration is 30%, and also fully precipitate it in an ice bath;- Centrifuge at 9000 rpm for 30 minutes at 10°C, discard the supernatant, and drain the pellet as dry as possible. Dissolve the precipitate in an appropriate amount of pH5.8, 0.01mol / LNa 2 HPO 4 -NaH 2 PO 4 buffer, aliquot. Dilute 5 ...

Embodiment 2

[0114] Take 200g clamworm, wash and mince, add 8 times the volume of pH8.0, 0.08mol / LTris buffer to make a homogenate; centrifuge at 6000 rpm for 60 minutes at 4°C, and take the supernatant, which is crude N-V protease Extraction. Take 1000ml of N-V protease crude extract, put it in an ice bath and cool it down to 0°C, add -20°C pre-cooled absolute ethanol under stirring conditions to make the concentration of ethanol reach 10%, keep it in an ice bath to make it precipitate ;-10°C, centrifuge at 6000 rpm for 60 minutes, discard the precipitate; add -20°C pre-cooled absolute ethanol to the supernatant while stirring, the final concentration is 25%, and also make it fully precipitate in an ice bath;- Centrifuge at 8,000 rpm for 30 minutes at 10°C, discard the supernatant, and drain the pellet as dry as possible. Dissolve the precipitate in an appropriate amount of pH 5.8, 0.01mol / L HAC-NaAC buffer, and distribute it. pH7.8, 0.04mol / L Na before use 2 HPO 4 -NaH 2 PO 4 The b...

Embodiment 3

[0116] Take 200g clamworm, wash and mince, add 8 times the volume of pH5.0, 0.04mol / LHAC-NaAC buffer to make a homogenate; centrifuge at 4000 rpm for 60 minutes at 4°C, take the supernatant N-V protease crude extract. Take 1000ml of N-V protease crude extract, put it in an ice bath and cool it down to 0°C, add -20°C pre-cooled absolute ethanol under stirring conditions to make the concentration of ethanol reach 10%, keep it in an ice bath to make it precipitate ;Centrifuge at -10°C, 4000 rpm for 60 minutes, discard the precipitate; add -20°C pre-cooled absolute ethanol to the supernatant while stirring, the final concentration is 25%, and also fully precipitate it in an ice bath;- Centrifuge at 10°C at 6000 rpm for 45 minutes, discard the supernatant, and drain the precipitate as much as possible. Dissolve the precipitate in an appropriate amount of pH 5.8, 0.01mol / L HAC-NaAC buffer, and distribute it. pH7.8, 0.04mol / L Na before use 2 HPO 4 -NaH 2 PO 4 The buffer solutio...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The present invention provides one kind of N-V proteinase and its separation and purification process. The N-V proteinase has log-in number in Swiss prot of P83433, molecular weight of 27000-35000, isoelectric point of 5-7.5, optimal temperature of 45 deg.c and optimal pH value of 7.8. Inside its molecule, the 10 peptide section amino acid residue sequence features (1) AVYLAGMK (2) NFPNYYINLY (3) VYLAANPTASS (4) QTFNSDTL and (5) VYILDTGI. The N-V proteinase is one kind of proteinase capable of hydrolyzing fibrinogen and fibrillarin powerfully both inside and outside body, and may be used as thrombolytic medicine and blood coagulation resisting medicine for preventing and treating myocardial infarction, cerebral artery thrombosis and other diseases.

Description

Technical field: [0001] The invention relates to a thrombolytic fibrinolytic enzyme, in particular provides a nereis protease and its separation, extraction and purification method and application, and relates to the biochemical technical field of the separation, extraction and purification method of the fibrinolytic enzyme. Background technique: [0002] Thromboembolic disease is the most common and serious blood circulation disorder in clinical practice. Treatment of the above diseases, especially the use of thrombolytic agents in the early stage of the disease, to reduce the content of fibrin and fibrinogen, reduce platelet aggregation, dissolve thrombus, and restore blood vessel recanalization, is the most important treatment method, which can reduce mortality and sequelae. At present, thrombolytic drugs such as urokinase, thromboxobinase and snake venom defibrase are commonly used clinically at home and abroad. These drugs are expensive and have different advantages an...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): A61K38/49C07K1/36C12N9/68
Inventor 洪敏程熠白若伦
Owner 洪敏
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com