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Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms

A Klebsiella, phage technology, applied in the direction of virus/phage, hydrolase, microorganism, etc., can solve problems such as difficult to remove

Active Publication Date: 2019-03-29
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, biofilm can be stably attached to the surface of medical devices, and it is difficult to remove by conventional physical and chemical means, which poses a severe challenge to the prevention and control of medical secondary infections

Method used

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  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms
  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms
  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms

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Embodiment 1

[0026] 1) The 32nd open reading frame from Klebsiella pneumoniae phage GH-K3 was cloned by means of PCR, double enzyme digestion and ligated molecular cloning. depo32 Gene ligation into pET28a plasmid: Restriction sites Nde I and xho I;

[0027] 2) The pET28a- depo32 The plasmid was transformed into Escherichia coli BL21(DE3), and BL21- depo32 Escherichia coli;

[0028] 3) Insert BL21- depo32 In 1L of LB liquid medium containing 50 μg / ml kanamycin, shake culture at 37°C until the logarithmic growth phase (OD 600 0.6~1.0);

[0029] 4) Add isopropyl-β-D-thiogalactoside to the culture medium to a final concentration of 1 mM, and shake at 16°C for 16 hours at 180 rpm;

[0030] 5) will induce the expression of BL21- depo32 After the bacterial liquid is collected by centrifugation, the bacteria are collected and centrifuged, and then ultrasonically crushed;

[0031] 6) Gently add the supernatant sample to the equilibrated Ni-NTA affinity chromatography column, was...

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Abstract

The invention discloses a bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms. By prokaryotic expression and purification of a 32nd open readingframe in a genome of Klebsiella pneumoniae bacteriophage GH-K3, a brand-new depolymerase depo32 with high activity is obtained. The depolymerase has high-efficiency degradation effect on the capsularpolysaccharides and biofilms formed by Klebsiella pneumoniae.

Description

technical field [0001] The invention discloses a phage depolymerase for degrading Klebsiella pneumoniae capsular polysaccharides and biofilms, relates to a method for degrading biofilms formed by Klebsiella pneumoniae by using phage depolymerization enzymes, and belongs to biotechnology field. Background technique [0002] Klebsiella pneumoniae ( Klebsiella pneumoniae ) is a common zoonotic pathogen. In recent years, multidrug-resistant Klebsiella pneumoniae has become one of the most important pathogens of hospital-acquired infections. The bacterium not only poses a huge threat to human public health, but also affects the healthy development of the animal breeding industry to a certain extent. [0003] Most Klebsiella pneumoniae have the ability to synthesize and secrete capsular polysaccharides. Capsular polysaccharides serve as a natural barrier for bacteria to maintain bacterial virulence, adherence, and block penetration of some antibiotics. As an important virulen...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N15/70C12N9/16A61K38/46A61P31/04
CPCA01N63/00A61K38/465A61P31/04C12N7/00C12N9/16C12N15/70C12Y301/00
Inventor 韩文瑜顾敬敏蔡若鹏程梦珺
Owner JILIN UNIV
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