A kind of separation and purification method of antibacterial protein
A technology for separation and purification of antimicrobial proteins, applied in peptide preparation methods, chemical instruments and methods, and methods based on microorganisms, can solve the problems of low yield of antimicrobial proteins, inability to obtain high-purity proteins, and many impurity proteins, etc., to achieve Good antibacterial activity, wide acid-base range, and strong stability
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Embodiment 1
[0034] (1) Preparation of B.amyloliquefaciens F1 fermentation broth
[0035]The B.amyloliquefaciens F1 bacterial strain was inoculated in a test tube containing 5mL of nutrient broth by 1% inoculum, and cultivated on a shaker at 33°C to obtain the B.amyloliquefaciens F1 seed liquid. Aspirate B.amyloliquefaciens F1 seed solution, add 2% inoculum amount to a Erlenmeyer flask with 40% loading capacity of fermentation medium, culture at 33°C on a shaker for 48 hours, and obtain B.amyloliquefaciens F1 fermentation solution.
[0036] (2) Extraction of B.amyloliquefaciens F1 antibacterial protein
[0037] First, take 1 L of the above-mentioned B. amyloliquefaciens F1 fermentation broth and perform ultrafiltration with an ultrafiltration membrane with a molecular weight cut-off of 15 kDa to obtain an ultrafiltration concentrate. Then slowly add ammonium sulfate to the ultrafiltration concentrate to make its saturation reach 0%, let it stand overnight at 4°C, and then centrifuge at 80...
Embodiment 2
[0041] (1) Preparation of B.amyloliquefaciens F1 fermentation broth
[0042] The B.amyloliquefaciens F1 bacterial strain was inoculated in a test tube containing 5mL of nutrient broth by 2% inoculation amount, and cultivated on a shaker at 30°C to obtain the B.amyloliquefaciens F1 seed liquid. Aspirate B.amyloliquefaciens F1 seed liquid, add 2% inoculum amount to an Erlenmeyer flask with 20% fermentation medium loading, and culture on a shaker at 32°C for 72 hours to obtain B.amyloliquefaciens F1 fermentation liquid.
[0043] (2) Extraction of B.amyloliquefaciens F1 antibacterial protein
[0044] First take 1L of the above-mentioned B.amyloliquefaciens F1 fermentation broth, and use an ultrafiltration membrane with a molecular weight cut-off of 8kDa to perform ultrafiltration to obtain an ultrafiltration concentrate. Then slowly add ammonium sulfate to the ultrafiltration concentrate to make its saturation reach 50%, let it stand overnight at 4°C, and then centrifuge at 10000...
Embodiment 3
[0055] (1) Preparation of B.amyloliquefaciens F1 fermentation broth
[0056] The B.amyloliquefaciens F1 bacterial strain was inoculated in a test tube containing 5mL of nutrient broth by 2% inoculation amount, and cultivated on a shaker at 30°C to obtain the B.amyloliquefaciens F1 seed liquid. Aspirate B.amyloliquefaciens F1 seed liquid, add 2% inoculum amount to an Erlenmeyer flask with 30% loading capacity of fermentation medium, and culture on a shaker at 37°C for 70 hours to obtain B.amyloliquefaciens F1 fermentation liquid.
[0057] (2) Extraction of B.amyloliquefaciens F1 antibacterial protein
[0058] First take 1L of the above-mentioned B.amyloliquefaciens F1 fermentation broth, and use an ultrafiltration membrane with a molecular weight cut-off of 15kDa to perform ultrafiltration to obtain an ultrafiltration concentrate. Then slowly add ammonium sulfate to the ultrafiltration concentrate to make the saturation reach 60%, let it stand overnight at 4°C, and then centri...
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