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Composite protective agent capable of improving stability of deep-sea elastase, and preparation method thereof
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An elastase and protective agent technology, which is applied in the directions of enzyme stabilization, biochemical equipment and methods, enzymes, etc., can solve the problems of easy inactivation, poor thermal stability, affecting development and application, etc., and achieves low cost and improved thermal stability. properties, improve the effect of raw materials
Active Publication Date: 2016-11-16
SHANDONG UNIV
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Problems solved by technology
As a result of long-term adaptation to the deep-sea low temperature environment, the thermal stability of the enzyme is very poor, and it is easily inactivated at lower temperatures, which seriously affects the development and application of the enzyme
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Embodiment 1
[0045] A compound protective agent for improving the stability of deep-sea elastase, the components per liter are as follows:
[0047] The preparation method of the composite protective agent for improving the stability of deep-sea elastase, the steps are as follows:
[0048] Add 371.57g of glucose, 472.91g of trehalose, 25mL of glycerin, and 85.57g of sucrose into 200ml of a solution with a concentration of 50mM Tris-HCl and pH9.0, stir evenly, and let stand at 65°C for 4 hours until the solute is completely dissolved, then add 50mM Tris-HCl HCl, pH 9.0 solution was adjusted to 1L. Sterilize under high temperature and high pressure at 115°C for 30 minutes, and store at room temperature for later use.
Embodiment 2
[0050] A compound protective agent for improving the stability of deep-sea elastase, the components per liter are as follows:
[0052] The preparation method is the same as in Example 1.
Embodiment 3
[0054] The preparation method of deep-sea elastase is as follows:
[0055] (1) Inoculate the deep-sea bacteria (Pseudoalteromonas sp.) CF6-2 with the strain preservation number CCTCC M 2010189 in the fermentation medium, culture at 20°C for 28 hours, and obtain the fermentation broth;
[0056] The fermentation medium component of described step (1) is as follows, is weight percent:
[0058] (2) Take the fermented liquid prepared in step (1), centrifuge at 10,000 g for 10 min at 5° C., take the supernatant, dialyze the supernatant with 50 mM Tris-HCl buffer solution of pH 9.0 overnight, and centrifuge at 10,000 g for 15 min at 5° C. , take the supernatant, after the supernatant is purified by DEAE-SepharoseFast Flowchromatography column, after passing through the DEAE-SepharoseFast Flowchromatography column at a speed of 2ml / min, gradient elution...
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Abstract
The invention relates to a composite protective agent capable of improving the stability of deep-sea elastase, and a preparation method thereof. The composite protective agent capable of improving the stability of deep-sea elastase is prepared from glucose, trehalose, glycerin, canesugar and a Tris-HCl solution and has a pH value of 8.0 to 9.0. The invention also relates to a preparation method for the composite protective agent. The composite protective agent provided by the invention can substantially improve the heat stability of an enzyme preparation; the composite protective agent is mixed with elastase liquid, about 80% of the activity of elastase can be maintained after preservation of the obtained mixture at 37 DEG C for three days while about 60% of the activity of elastase can be maintained after preservation for five days, and 90% of the activity of elastase can be maintained after preservation of the mixture at 4 DEG C for seven months; so the problem of proneness to inactivation of deep-sea elastase after long-term preservation is overcome. The composite protective agent can substantially improve the heat stability of a plurality of enzymes.
Description
technical field [0001] The invention relates to a compound protective agent for improving the stability of deep-sea elastase and a preparation method thereof, belonging to the technical field of biotechnology. Background technique [0002] Elastase is a kind of proteolytic enzyme that can hydrolyze insoluble elastin. Its substrate elastin is mainly distributed in connective tissues such as blood vessels, lungs and skin. Therefore, elastase can be used to treat hyperlipidemia, prevent arterial Biochemical drugs for atherosclerosis, elimination of skineschar, debris, etc. Because elastase can specifically degrade the tough elastic fiber components in connective tissue, elastase can be used as an ideal meat tenderizer in the food processing industry. Due to the similarity in the partial sequence of the peptides of elastin and gram-positive bacterial peptidoglycan, elastase can sometimes be used as lysozyme. Lysozyme can be used as a natural foodpreservative. It has been wid...
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