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Spherical bacillus and organic solvent-resistant stable proteinase produced thereby

A spheroid bacillus, organic solvent technology, applied in the directions of bacteria, hydrolase, microorganisms, etc., can solve the problems of no industrial production, no spheroid bacillus, difficult to meet the needs of the market, etc.

Inactive Publication Date: 2009-04-15
HUAIHAI INST OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

And there is no industrial production, it is difficult to meet the needs of the market
[0004] There are few domestic research reports on organic solvent-resistant microorganisms and organic solvent enzyme stability. At present, there is no report on organic solvent-stable protease produced by Bacillus sphaericus in China.

Method used

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  • Spherical bacillus and organic solvent-resistant stable proteinase produced thereby
  • Spherical bacillus and organic solvent-resistant stable proteinase produced thereby
  • Spherical bacillus and organic solvent-resistant stable proteinase produced thereby

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1. Isolation and cultivation of Bacillus sphaericus DS11 strain.

[0050] Add 1ml of seawater sample taken from the Lianyungang waters of the Yellow Sea, China, into a 250ml Erlenmeyer flask filled with 50mL of MLB liquid medium containing 10% benzene, plug the bottle mouth with a rubber stopper, and transfer to In the new MLB liquid medium, if it is 3 times; Dilute the obtained culture solution by 10 5 After doubling, draw 10 μl of coated MLB medium plate, culture in 28°C incubator for 2 days, observe the colony shape, streak and purify to pure species, then plant on SMA medium plate, and culture in 28°C incubator for 2 days, preliminary Screen out the strains whose ratio of the colony diameter to the diameter of the transparent circle is greater than 5, and inoculate the primary screened strains into the enzyme-producing fermentation medium, culture at 28°C and 180r / min for 48h, centrifuge at 8000g for 10min at 4°C, and measure the supernatant Protease activ...

Embodiment 2

[0051] Example 2. Characteristics of Bacillus sphaericus DS11 strain.

[0052] The strain is Gram-positive, short rod-shaped, 2.2-2.7 μm×0.7-1.3 μm in size, non-capsulated, and spore-forming. The colonies on the solid medium of skimmed milk (SMA) are 2mm-4mm in diameter, white, moist, with smooth edges, protruding in the middle, easy to pick, and produce obvious transparent circles. The catalase, urease, and MR tests of the bacteria are all positive, and the oxidase and VP tests are negative, and cannot produce indole and H2S; it can hydrolyze gel, Tween-80, triolein, and casein , can not hydrolyze starch. Strain DS11 can utilize glucose, sucrose, xylose, and mannitol, but cannot utilize maltose, lactose, raffinose, rhamnose, and malonic acid as carbon sources.

Embodiment 3

[0053] Example 3. Study on resistance to organic solvents of Bacillus sphaericus DS11 strain.

[0054] Inoculate the Bacillus sphaericus DS11 described in Example 1 onto the MLB solid medium plate, cover the organic solvent with 7.0ml of different polar constants on the plate, seal it with a plastic wrap to prevent the organic solvent from volatilizing, and cultivate it at 28°C for 48h. The bacterial strain DS11 can Grow on a plate covered with an organic solvent with a polarity constant not greater than 2 to form colonies.

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PUM

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Abstract

The invention discloses a bacillus sphaericus DS 11. The strain is a short rod shape Gram-positive bacteria, the growth temperature range of the strain is 4 to 35 DEG C and the optimal growth temperature is 28 DEG C; the growth pH range is between 4 and 11, and the optimal growth pH is 7; the growth NaCl concentration is between 0.3 and 10 percent, and the optimal NaCl concentration is 3 percent. The bacillus sphaericus DS 11 can endure an organic solvent with Log P of less than or equal to 2.0. The invention further discloses a method for producing an inactive protease of the organic solvent by utilizing the bacillus sphaericus DS 11 as well as the inactive protease product of the organic solvent which is obtained by the method, has stronger organic solvent tolerance and can be used for the synthesis of organic-phase high-performance catalytic polypeptide.

Description

technical field [0001] The invention relates to a kind of bacillus sphaericus DS11 (Bacillus sphaericus DS11) isolated from the seawater of Lianyungang sea area in the Yellow Sea of ​​China; the invention also relates to a method and a product of the organic solvent-stable protease produced by the bacillus sphaericus. Background technique [0002] Protease can catalyze the hydrolysis of peptide bonds in the aqueous phase, and catalyze the synthesis of peptide bonds in the organic phase. It is one of the most widely used enzyme preparations and is widely used in peptide synthesis, protein processing, food, pharmaceuticals, leather and detergents and other fields. When proteases catalyze peptide synthesis in organic phases, proteases are required to be resistant to organic solvents. However, organic solvents are highly toxic to most enzymes. The tolerance of enzymes to organic solvents can be improved by means of chemical modification, embedding, immobilization, protein engi...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/54C12R1/07
Inventor 吕明生房耀维王淑军刘姝朱广超
Owner HUAIHAI INST OF TECH
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