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Bacterial strain for producing low temperature alkaline protease

A protease and bacterial strain technology, applied in the direction of bacteria, hydrolytic enzymes, and microbial-based methods, can solve the problems of low catalytic efficiency and no low-temperature basic protein production, and achieve the effect of excellent properties and high protease activity

Active Publication Date: 2015-09-30
QINGDAO AGRI UNIV
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  • Abstract
  • Description
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AI Technical Summary

Problems solved by technology

At present, the alkaline proteases used as detergent additives are mostly medium-temperature proteases, and the optimum reaction temperature is usually 50-70°C, but in developing countries, detergents are usually used at room temperature around 25°C, so the alkaline protease added to the detergent The catalytic efficiency of protease is not high, many researchers began to study low temperature alkaline protease
However, there is no practically applied strain producing low-temperature basic protein

Method used

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  • Bacterial strain for producing low temperature alkaline protease
  • Bacterial strain for producing low temperature alkaline protease
  • Bacterial strain for producing low temperature alkaline protease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1: Screening and identification of bacterial strains

[0029] Separation of endophytic bacteria from wakame seaweed collected in Qingdao sea area, and plant the obtained single bacterial colony on the casein medium, observe the situation of each strain producing hyaline circles, and carry out preliminary screening of bacteria species; then produce hyaline circles The strain was inoculated on the liquid fermentation medium, cultured in a shake flask at 25°C for 3 days, centrifuged at 10,000 r / min at 4°C for 15 minutes, and the supernatant was taken to measure its protease activity, and the strain with the highest protease activity was screened out.

[0030] According to "Berger's Bacterial Identification Handbook" and "Systematic Identification", the WFWBac-1 strain was classified and identified according to its physiological and biochemical characteristics.

[0031] Physiological and biochemical identification tests were carried out on the WFWBac-1 strain. Th...

Embodiment 2

[0056] The fermentation condition optimization of embodiment 2 bacterial strains

[0057] 1. Effect of carbon source on enzyme production by WFWBac-1 bacteria

[0058] The protease activity was determined after the carbon source to be tested (glucose, sucrose, starch, fructose, lactose) was used as the sole carbon source in the liquid medium, cultured at 25°C and 150r / min for 48h. The result is as figure 1 It was shown that when sucrose was used as carbon source, the enzyme activity of WFWBac-1 strain was the highest, glucose, lactose, starch were lower, and fructose was the lowest. It can be seen that the strain can make full use of the carbon source in sucrose for growth, metabolism and reproduction, so sucrose is the best carbon source for WFWBac-1.

[0059] 2. Effect of nitrogen source on enzyme production by WFWBac-1 bacteria

[0060]In the nitrogen source test, sucrose was used as a fixed carbon source; the nitrogen source to be tested was casein, sodium nitrate, ammo...

Embodiment 3

[0069] The research of the crude enzyme liquid property of embodiment 3 bacterial strains

[0070] 1. Determination of the optimum reaction temperature of the enzyme

[0071] Put the crude enzyme solution produced by the fermentation of the screened strain WFWBac-1 at 0, 20, 40, and 80°C respectively for 1 hour, take out the crude enzyme solution, add the substrate azo casein and place it at 37°C , using the Folin-phenol method to measure the enzyme activity and determine the optimum reaction temperature of the enzyme. Such as Figure 7 As shown, the optimum temperature of protease in the crude enzyme solution is 20°C, and the enzyme activity is 1078U / mL at this time, which is the result of the comprehensive action of various proteases, indicating that the protease produced by this bacteria belongs to low-temperature protease.

[0072] 2. Determination of the optimum pH value of the enzyme

[0073] The crude enzyme solution produced by the fermentation of the screened strai...

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Abstract

The invention provides a bacterial strain for producing low temperature alkaline protease. The bacterial strain is Bacillus sp. WFWBac-1 and is preserved at a general microbiological center of a China committee for culture collection of microorganisms of the institute of microbiology of Chinese academy of sciences located at courtyard 1, No 3, Beichen west road, Chaoyang District of Beijing on April 9, 2015, and the preservation number is CGMCC NO.10703. The filtered bacterial strain is used for producing the alkaline protease in a fermentation mode and is good in performance and high in vitality of produced protease. The carbon source of the bacterial strain is cane sugar, the nitrogen source of the bacterial strain is peptone, the initial pH value of a culture medium is 9.0, the culture temperature is 25 DEG C, and when the bacterial strain is shaken and cultured for 48 h at 150 rpm under the condition of 10% of the inoculum size, the vitality of the produced protease is as high as 995 U / mL. The prepared protease is used for preparing liquid detergents or cleanser essences.

Description

technical field [0001] The invention belongs to the technical field of functional microorganism screening, and in particular relates to a strain producing low-temperature alkaline protease. Background technique [0002] Protease is a very important hydrolytic enzyme. Relevant data at home and abroad show that in all industrial enzyme preparations, protease accounts for 75%, which is the largest proportion of industrial enzymes, and its sales account for about 100% of the world's total annual sales. About 60%. Alkaline protease is one of the most important enzyme preparations in the world's industrial enzyme preparations. It has a wide range of uses and has attracted widespread attention. At present, alkaline protease is mainly used in washing and leather industries, among which more than 99% of detergents are more or less added with various proteases, resulting in a demand relationship that is in short supply today. At present, the alkaline proteases used as detergent addi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/54C11D3/386C12R1/07
Inventor 王凤舞
Owner QINGDAO AGRI UNIV
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