Preparation and characterization of recombinant human-source collagen.
- Author:
Zengmiao HOU
1
;
Xiaoying LI
1
;
Min LI
1
;
Jinfang YANG
1
;
Xiaolin YANG
1
;
Jinli ZHAO
1
Author Information
- Publication Type:Journal Article
- Keywords: Pichia pastoris; biomaterial; collagen; human-source
- MeSH: Amino Acid Sequence; Biocompatible Materials; Collagen; analysis; Freeze Drying; Humans; Pichia; Recombinant Proteins
- From: Chinese Journal of Biotechnology 2019;35(2):319-326
- CountryChina
- Language:Chinese
- Abstract: This study aimed to obtain a recombinant human-source collagen for industrialization. First, based on the Gly-X-Y sequence of human type I collagen, we optimized the hydrophilic Gly-X-Y collagen peptide, designed the human collagen amino acid sequence and the corresponding nucleotide sequence. Next, the expression vector pPIC9K-COL was constructed via endonuclease digestion technology. We obtained an engineering strain of human-source collagen by electrotransforming Pichia pastoris, and then it was fermented, purified and identified. As a result, the expression level reached 4.5 g/L and the purity was over 95%. After amino acid N-terminal sequencing, molecular weight analysis, amino acid analysis and collagenase degradation test, we confirmed that the obtained collagen was consistent with designed primary structure of human-source collagen. After freeze-drying, we analyzed the collagen by scanning electron microscope and cell cytotoxicity, confirming that the collagen has porous fiber reticular structure and superior cytocompatibility. This indicates that human-source collagen has potential to be applied as biomedical material. In conclusion, we successfully obtained the expected human-source collagen and laid a foundation to its further application.
