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Streptomyces aureus producing keratinase and its application method

A technology of Streptomyces aureus and keratinase, applied in the field of microorganisms, can solve problems such as not easy to be hydrolyzed by ordinary proteases, unused, and environmental pollution

Active Publication Date: 2015-09-30
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because keratin has a large number of disulfide bonds, hydrogen bonds and hydrophobic interactions, it is not easy to be hydrolyzed by common proteases, so most of them are not used, and some even cause environmental pollution.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Example 1 Screening and identification of keratinase-producing strains

[0015] 1. Culture medium formula

[0016] (1) Primary screening medium (g / l): wool powder 5, K 2 HPO 4 1, NaCl 0.5, agar powder 20.

[0017] (2) Seed liquid culture medium

[0018] Gao's medium (g / l): soluble starch 20, KNO 3 1, NaCl 0.5, K 2 HPO 4 0.5, MgSO 4 . 7H 2 O 0.5,

[0019] FeSO 4 7H 2 O 0.01.

[0020] LB medium (g / l): peptone 10, yeast powder 5, sodium chloride 10.

[0021] Cha's medium (g / l): sucrose 30, NaNO 3 3,K 2 HPO 4 1, MgSO 4 . 7H 2 O 0.5, KCl 0.5, FeSO 4 . 7H 2 o

[0022] 0.01.

[0023] (3) Fermentation medium (g / l): wool powder 10, K 2 HPO 4 0.4, NaCl 0.5

[0024] 2. Plate preliminary screening and determination of keratin activity

[0025] Soil samples were collected from sheep pens in rural areas near Jiangnan University, and 1 g of soil samples were added to a 50 ml Erlenmeyer flask containing 10 ml of normal saline and a few glass beads, s...

Embodiment 2

[0031] Example 2 Preliminary Optimization of Strain Fermentation Conditions

[0032] The fermentation medium and fermentation conditions of the strain were optimized by single factor and orthogonal experiments, and the optimum medium formula was 0.5-1% wool, 4-8% glucose, 3-5% peptone, 0.3% sodium nitrate, phosphoric acid Dipotassium hydrogen dipotassium 0.04%, sodium chloride 0.05%, shaker culture conditions: initial pH value 10.0, inoculum size 10%, culture temperature 30°C, shaker speed 220r / min, liquid volume 30ml / 250ml, fermentation cycle 64h. Its keratinase activity reaches 44.2U / ml, which is 10.3 times higher than before optimization.

Embodiment 3

[0033] Example 3 Research on properties of crude enzyme

[0034] The crude enzyme properties of the produced keratinase were preliminarily studied, and the effects of the action temperature (30, 40, 45, 50, 55, 60, 70 and 80°C) and the action pH (3.0-10.0) on the enzyme activity were studied respectively. It was found that the optimum action temperature of the keratinase was 50-60°C, the optimum pH was 8.5, and the enzyme activity was stable (±10%) in an alkaline (pH7.0-10.0) environment. The enzyme is a mesophilic alkaline keratinase.

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PUM

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Abstract

The invention discloses a keratin producing bacterial strain as well as culture conditions and crude enzyme property thereof and belongs to the technical field of microorganisms. Through identification, the bacterial strain is Streptomyces aureofaciens, named as K13 and preserved in CGMCC (China General Microbiological Culture Collection Center) with the preservation number of CGMCC NO.8047. Through preliminary optimization, the culture medium of the bacterial strain comprises 0.5-1% of wool, 4-8% of glucose, 3-5% of peptone, 0.3% of sodium nitrate, 0.04% of dipotassium phosphate and 0.05% of sodium chloride; the shake culture conditions are that the initial pH (potential of Hydrogen) value is 10.0, the inoculation quantity is 10%, the culture temperature is 30 DEG C, the rotating speed is 220 revolutions per minute, the liquid loading volume is 30ml / 250ml, the fermentation period is 64 hours and the keratinase producing activity is up to 44.2U / ml. The keratinase is mesophilic alkaline keratinase, the optimum operation temperature is 50-60 DEG C and the optimum pH is 8.5; the enzyme activity of the keratinase is stable in an alkaline environment (pH 7.0-10.0).

Description

technical field [0001] The invention belongs to the technical field of microorganisms, and in particular relates to a strain of Streptomyces aureus producing keratinase and properties of crude enzymes thereof. Background technique [0002] my country produces a large amount of poultry feathers and waste wool every year, the main component of which is keratin. Because keratin has a large number of disulfide bonds, hydrogen bonds and hydrophobic interactions, it is not easy to be hydrolyzed by common proteases, so most of them are not used, and some even cause environmental pollution. Keratinase can degrade poultry feathers and waste wool into soluble proteins, polypeptides or amino acids, which can be used as medical raw materials, animal feed, etc., turning waste into treasure and reducing environmental pollution. Many microorganisms can produce keratinase, such as Streptomyces, Monas, Bacillus and so on. [0003] The purpose of the present invention is to provide a strai...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N9/52C12R1/49
Inventor 张旦旦王月史劲松许正宏张晓梅
Owner JIANGNAN UNIV
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