Method for expressing pseudoplectania nigrella mature peptide in recombinant pichia pastoris

Abstract

The invention provides a method for expressing and producing pseudoplectania nigrella mature peptide recombinant proteins in pichia pastoris cells in a constitutive type mode, which comprises the following steps of: 1) optimizing pseudoplectania nigrella mature peptide genes; 2) obtaining pichia pastoris glyceraldehyde-3-phosphate dehydrogenase promoter DNA sequences by adopting methods such as molecular cloning and the like, and placing the pseudoplectania nigrella mature peptide genes under the regulation and control of the glyceraldehyde-3-phosphate dehydrogenase promoters to connect with plasmids to obtain new, constitutive and secretory type pichia pastoris efficient expression vectors; 3) converting the pichia pastoris by using the vectors and obtaining the recombinant pichia pastoris strains to obtain the pseudoplectania nigrella mature peptide with bioactivity; and 4) optimizing the high-density fermentation conditions of the engineering strains and the expression and purification of the recombinant protein. The pseudoplectania nigrella mature peptide prepared by the method can be applied in the fields of medicinal treatment, food, feed, scientific research and the like and is suitable for the scale production of pseudoplectania nigrella protein.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a method for expressing a mature pseudosantagactin polypeptide in recombinant Pichia pastoris. Background technique [0002] Antimicrobial peptides (AMPs) is a general term for a class of small molecule polypeptides composed of 12-100 amino acids, with a net positive charge, amphipathic and antibacterial activity. The research on antimicrobial peptides has a history of more than 50 years. So far, about 880 antimicrobial peptides from various biological sources have been identified (Brogden K A, 2005). Antimicrobial peptides have attracted increasing attention due to their broad-spectrum antibacterial activity (including against Gram-positive bacteria, Gram-negative bacteria, fungi, and even some viruses, etc.). Especially in the past few decades, due to the abuse of antibiotics, pathogenic strains with antibiotic resistance have emerged continuously, and there is an urgent nee...

AI Technical Summary

Problems solved by technology

[0011] At present, the expression of exogenous proteins in Pichia pastoris is generally through the integration of exogenous genes into the Pichia genome, although there are also some expression plasmids (such as 2μ) carrying foreign genes that can replicate independently in Pichia pastoris and There are reports on the expression of foreign proteins, but this non-integrated expression method has disadvantages such as plasmid instability and easy loss, and is rarely used at present.

Images