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Recombinant human papillomavirus type 6 l1 protein and its use

A protein, pentamer technology, applied in the direction of viruses, viral peptides, antiviral agents, etc., can solve the problems of inability to produce protective antibodies, inability to apply large-scale production, and difficulty of target proteins.

Active Publication Date: 2018-10-19
BEIJING HEALTH GUARD BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, because most of the HPV L1 protein expressed by E. coli loses its natural conformation, it cannot produce protective antibodies against HPV.
Or although the above protein can be purified by inclusion bodies, renaturation and other steps can also obtain HPV VLP, but the loss of protein is large during the renaturation process, and the yield is low, so it is difficult to apply in large-scale production.
Although the full-length sequence protein of HPV L1 can also be solublely expressed in the correct conformation in Escherichia coli and dissolved in the lysed supernatant of the bacteria, the expression level is low, and there are many types and large amounts of foreign proteins in the supernatant. It is quite difficult to purify the target protein, and it is still not applicable to large-scale production

Method used

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  • Recombinant human papillomavirus type 6 l1 protein and its use
  • Recombinant human papillomavirus type 6 l1 protein and its use
  • Recombinant human papillomavirus type 6 l1 protein and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment l

[0067] Embodiment 1: the construction of the engineering bacteria with HPV6 L1 sequence 2

[0068] 1. The full-length HPV6 L1 gene was synthesized by GENEWIZ, whose gene sequence is SEQ ID NO: 1, which is derived from GeneBank, and the sequence number is NC_001355.1. aa

[0069] 2. The template containing the gene fragment of SEQ ID NO: 1 for PCR reaction. Use the forward primer sequence: 5'-CGCGGATCCGGA AAACCGGACCCGTACAAAAAC-3'; introduce the restriction endonuclease AccIII site at the 5' end of the H4 structure, and the AccIII site sequence is TCCGGA. The downstream contains the XhoI endonuclease site, the reverse primer sequence: 5'-GCTCTCCTCGAGAACACCGGTA CGGATAGAAG A-3', the 5' end introduces the restriction endonuclease XhoI site, and the XhoI site sequence is CTCGAG. HPV6B was obtained by PCR amplification.

[0070]3. A template containing the gene fragment of SEQ ID NO: 1 for PCR reaction. To contain the introduced restriction endonuclease BamHI site, the BamH I sit...

Embodiment 2

[0075] Example 2: Construction of engineering bacteria with HPV6 L1 sequence 4

[0076] 1. The full-length target gene fragment of HPV6 L1 gene was purchased from the clinical cell sample waste containing wild-type HPV6 virus in the gynecological clinic of Beijing Anzhen Hospital, and its gene sequence is SEQ ID NO: 3 (AF067044.1).

[0077] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. Use the forward primer sequence: 5'-CGCGGATCCGGA AAGCCAGATCCCTATAAGAAC-3'; introduce a restriction endonuclease AccIII site at the 5' end of the H4 structure, and the AccIII site sequence is TCCGGA. The downstream contains the XhoI endonuclease site, the reverse primer sequence: 5'-GCTCTCCTCGAGTTAGGCAGCA GAGGCTCTGG AAAC-3', the 5' end introduces the restriction endonuclease XhoI site, and the XhoI site sequence is CTCGAG. After PCR reaction, HPV6B was obtained by amplification.

[0078] 3. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. ...

Embodiment 3

[0083] Example 3: Construction of engineering bacteria with HPV6 L1 sequence 6

[0084] 1. The full-length HPV6 L1 gene was synthesized by GENEWIZ, whose gene sequence is SEQ ID NO: 5, which is derived from GeneBank, and the sequence number is HE962027.1.

[0085] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. Use the forward primer sequence: 5'-CGCGGATCCGGA AAGCCAGATCCCTATAAGAAC-3'; introduce a restriction endonuclease AccIII site at the 5' end of the H4 structure, and the AccIII site sequence is TCCGGA. The downstream contains the XhoI endonuclease site, the reverse primer sequence: 5'-GCTCTCCTCGAGTTAGGCAGCAGAG GCTTTGGAAA C-3', and the restriction endonuclease XhoI site is introduced at the 5' end, and the XhoI site sequence is CTCGAG. PCR reaction, amplification to obtain HPV6B.

[0086] 3. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. The forward primer sequence containing the introduced restriction endonuclease ...

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Abstract

The invention provides a novel polynucleotide gene segment of coded recombinant protein HPV6L1, a vector containing the gene segment, a host cell containing the vector, a protein HPV6L1 pentamer obtained by translating and expressing the gene segment, and an HPV6 infection-resistant condyloma acuminate vaccine consisting of the pentame.

Description

technical field [0001] The present invention relates to the prevention and / or treatment of human papillomavirus infection. More specifically, the present invention relates to a recombinant human papillomavirus type 6 L1 protein, a pentamer composed thereof, a vaccine containing the protein and a vaccine for preventing HPV6 type virus infection, especially in the prevention of HPV6 type. Use in condyloma acuminatum disease caused by viral infection. Background technique [0002] Human papillomavirus (Human Papillomavirus, referred to as HPV) is a DNA virus transmitted through close contact. In human tissues, HPV mainly infects skin and mucosal tissues. HPV can be divided into high-risk and low-risk types. Long-term persistent infection of high-risk types can cause cancer, such as cervical cancer, which is life-threatening. Low-risk HPV can cause genital lesions, such as condyloma acuminatum, affecting quality of life. Condyloma acuminatum is the most common sexually trans...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/025C12N15/37C12N15/63C12N5/10C12N1/15C12N1/19C12N1/21C07K19/00A61K39/12A61P31/20
CPCA61K39/00C07K14/005C12N2710/20022C12N2710/20034
Inventor 刘永江陈小江陈林盖大海许铮曹科陈建平潘勇昭银飞阮芳勇
Owner BEIJING HEALTH GUARD BIOTECH