A kind of recombinant human type III collagen and its Pichia pastoris recombinant expression system

A collagen and repeat sequence technology, applied in the field of genetic engineering, can solve problems affecting protein immunogenicity and protein application, and achieve the effects of rapid production method, good biocompatibility and high stability

Active Publication Date: 2022-02-01
西安德诺海思医疗科技有限公司
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Problems solved by technology

At the same time, although the target protein with higher purity can be obtained by using specific purification tags, the purification tags will affect the immunogenicity of the protein, thereby affecting the application of the protein

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  • A kind of recombinant human type III collagen and its Pichia pastoris recombinant expression system
  • A kind of recombinant human type III collagen and its Pichia pastoris recombinant expression system
  • A kind of recombinant human type III collagen and its Pichia pastoris recombinant expression system

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Embodiment Construction

[0033] The invention will be described in further detail below in conjunction with the accompanying drawings and embodiments, which are only used to explain the present invention rather than limit the protection scope of the present invention.

[0034] (1) Structural design of human collagen

[0035] According to the amino acid sequence of natural human type III collagen (reference sequence Genebank accession number: NM_000090.3), the present invention selects a peptide segment consisting of 169 amino acids, and the peptide segment contains valine (V), phenylpropanoid The Gly-X-Y structural peptides of amino acid (F), leucine (L), methionine (M) or alanine (A) are all optimized. The main optimization method is: using natural human type III collagen The helical region is replaced by a Gly-X-Y structural peptide that does not contain the above amino acids with a high frequency (≥3 times).

[0036] The 169AA sequence (position 537-705 in the reference sequence) selected by the p...

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Abstract

The invention discloses a recombinant human type III collagen and a Pichia pastoris recombinant expression system thereof. In the present invention, the Gly-X-Y structural peptides containing valine, phenylalanine, leucine, methionine, and alanine in the partial peptides of human type III collagen are used as the peptides with higher frequency of use. The Gly-X-Y structure peptide containing the above amino acids is replaced, and the gene sequence encoding the peptide is obtained according to the peptide after replacement, and then a gene expression vector for tandem expression of peptides with different repeat numbers is constructed and transformed into a Pichia host strain, screened A strain highly expressing recombinant human type III collagen was obtained. The invention effectively solves the problem of degradation of natural collagen during fermentation through the artificial design and optimization of the protein structure, and the expression product has high stability and good biocompatibility, and can be used for large-scale production.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to the expression of recombinant human type III collagen by Pichia pastoris. Background technique [0002] Collagen is the most abundant protein in the body and the main component of the extracellular matrix (ECM). It plays an important role in maintaining the normal physiological functions and repairing damage of cells, tissues and organs. It has been widely used in medicine, Health care products and cosmetics industry. In terms of molecular structure, each collagen peptide chain is mainly composed of Gly-X-Y structural peptides (X, Y are any amino acid residues other than Gly), this peptide chain structure is necessary for the formation of collagen fiber advanced structure, determines Collagen has excellent biocompatibility and low immunogenicity. [0003] The main source of collagen is extracted from animal tissues. With the development of biotechnology, recombina...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/78C12N15/81C12N15/12C12N1/19A61L27/24A61K8/65A61Q19/00C12R1/84
CPCC07K14/78C12N15/815A61L27/24A61K8/65A61Q19/00A61K2800/10A61K2800/805
Inventor 杨鹭黄文涛
Owner 西安德诺海思医疗科技有限公司
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