Anthrax resisting polypeptide and its application and preparation method

An anti-anthrax and anthrax rod technology, applied in the direction of botany equipment and methods, biochemical equipment and methods, applications, etc., can solve problems such as not very effective

Active Publication Date: 2008-07-09
PROTEIN DESIGN LAB LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the above-mentioned means are not very effective, human beings

Method used

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  • Anthrax resisting polypeptide and its application and preparation method
  • Anthrax resisting polypeptide and its application and preparation method
  • Anthrax resisting polypeptide and its application and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1 Construction of a plasmid expressing an anti-anthrax polypeptide and preparation of a recombinant anti-anthrax polypeptide

[0019] The original plasmid is the pET22b plasmid (7.6kb, the blank plasmid is the Novagan pET22b plasmid, and the PA gene is loaded by Harvard Univ.Dr.J.Collier) loaded with the wild-type Bacillus anthracis protein antigen (PA) gene SEQ ID NO.1. Chain oligonucleotide point mutation technology (QuickChange TM Kit, Strategene Company) after inserting the gene-CACCACCACCACCACCAC- of six histidines at the amino terminus of the PA gene, the genes (SEQ ID NO. The bacillin antigen (PA) gene is operably linked to obtain the nucleotide sequence SEQ ID NO.10 encoding the recombinant mutant bacillin antigen gene. After inserting the gene SEQ ID NO.12 encoding the antibody mimic into the carboxy-terminal G735 site of the recombinant mutant Bacillus anthracis protein antigen gene, the recombinant plasmid pCHCA-PA1 of the anti-anthrax polypeptide w...

Embodiment 2

[0075] Example 2: The protective effect of the anti-anthrax polypeptide of the present invention on mice killed by the wild-type Bacillus anthracis PA / LF complex

[0076] The tested mice were BALB / c immunodeficient nude mice, which were divided into 4 test groups, each with 10 mice. Wild-type Bacillus anthracis PA / LF complex, anti-anthrax polypeptide, and anti-tumor binary polypeptide were administered intraperitoneally, once a day at the same time, for a total of 6 times, and the dead mice were no longer administered.

[0077] Experimental group 1: intraperitoneal injection of wild-type Bacillus anthracis PA / LF complex group, the injection volume was 20 μg / 20 μg / day.

[0078] Experimental group 2: intraperitoneal injection of anti-anthrax polypeptide / LF complex group, the injection volume was 20 μg / 20 μg / day.

[0079] Experimental group 3: Intraperitoneal injection of wild-type Bacillus anthracis PA / LF complex + anti-anthrax polypeptide group, the injection volume was 20 μg / ...

Embodiment 3

[0082] Example 3: The protective effect of the anti-anthrax polypeptide of the present invention on mice killed by the wild-type Bacillus anthracis PA / LF complex after a delay of 6 hours

[0083] The tested mice were BALB / c immunodeficient nude mice, which were divided into two test groups, each with 5 mice. The wild-type Bacillus anthracis PA / LF complex and the anti-anthrax polypeptide were administered intraperitoneally, and the anti-anthrax polypeptide was administered 6 hours after the wild-type Bacillus anthracis PA / LF complex was administered on the first day. Thereafter administered once daily at the same time. A total of 6 doses were administered, and the dead mice were no longer administered.

[0084] Experimental group 1: intraperitoneal injection of wild-type Bacillus anthracis PA / LF complex group, the injection volume was 20 μg / 20 μg / day.

[0085] Experimental group 2: Intraperitoneal injection of wild-type Bacillus anthracis PA / LF complex + anti-anthrax polypept...

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Abstract

The invention provides an anti-anthrax polypeptide gene, a recombinant plasmid, a polypeptide, the application and a process for preparation which comprises following steps: operationally connecting a gene of an encoding antibody simulacrum with the gene of an encoding recombinant mutation bacillus anthracis proteantigen to obtain the gene which expresses a recombinant anti-anthrax polypeptide, inserting the gene of the encoding antibody simulacrum into the gene of the recombinant mutation bacillus anthracis proteantigen through the technique of double chain oligonucleotide point mutation to form the recombinant plasmid of the invention, transfecting the recombinant plasmid which is obtained into coli bacteria BL-21 project engineering bacteria to obtain engineering bacteria cells of the anti-anthrax polypeptide, obtaining the anti-anthrax polypeptide through extracting supernate which contains polypeptide from a great amount of increasing bacteria, centrifugal precipitation cells and saccharose with an addex-magnesiumand method and purifying the supernate with florisil column. The anti-anthrax polypeptide specifically can damage the biological activity of bacillus anthracis toxin and does not attack normal human cells.

Description

technical field [0001] The invention relates to an anti-anthrax polypeptide gene, recombinant plasmid, polypeptide and application and preparation method thereof. The anthrax is a lethal infectious disease caused by Bacillus anthracis or anthrax toxin. Background technique [0002] Fatal infectious diseases caused by Bacillus anthracis or anthrax toxin have always been a huge threat to human health. In addition, Bacillus anthrax or anthrax toxin has been the most feared pathogen and toxin in bioterrorism attacks. For fatal infectious diseases caused by Bacillus anthracis or anthrax toxin, the current prevention and treatment methods mainly use antibiotics, antivenom, antibody therapy and vaccination. Since the above means are not very effective, human beings have been actively looking for new ways of prevention and treatment. Contents of the invention [0003] One object of the present invention is to provide an anti-anthrax polypeptide gene, recombinant plasmid, and pol...

Claims

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Application Information

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IPC IPC(8): C12N15/31C12N15/13C12N15/63C12N15/70C07K14/195A61K39/02A61K39/395A61P31/04
Inventor 丘小庆
Owner PROTEIN DESIGN LAB LTD
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