Method for realizing efficient covering of foreign protein by using insect virus polyhedron

Abstract

The invention discloses a method for realizing efficient covering of a foreign protein by using an insect virus polyhedron, wherein the polyhedrin of a bombyx mori cytoplasmic polyhedrosis virus (BmCPV) and a target protein gene are constructed under the PH and P10 promoters on a pFastBac Dual carrier, and a baculovirus capable of expressing the polyhedron and the target protein simultaneously can be obtained through recombination. The method is characterized in that the cytoplasmic polyhedrosis protein gene and the target protein gene are simultaneously constructed under the PH and P10 promoters on the pFastBac TMDual carrier through a baculovirus expression system, respectively, a flexible link peptide is introduced between the target protein and a signal peptide, and the recombinant baculovirus is obtained through recombination with a virus skeleton. Once Sf9 or Sf21 insect cells are infected by the single virtus, the desired covered protein can be directly generated in cells; the recombinant virtus also can be amplified.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a method for wrapping proteins. Background technique [0002] Insect virus polyhedron (polyhedra) refers to the polyhedral particles produced by insect viruses after infecting host cells, which are mainly used to wrap virus particles. Its function is to protect the encapsulated virus particles from adverse environmental damage, such as: high temperature, dryness, radiation, etc., and it is also resistant to various proteolytic enzymes, and is extremely stable in acidic conditions, while in alkaline environments It dissolves easily and releases virus particles [1]. Polyhedrons can be classified as nuclear polyhedrons or cytoplasmic polyhedrons according to the location of polyhedron formation in the cell. Recent studies at home and abroad have found that plastid polyhedrons can wrap heterologous proteins. For example, Ijiri et al. have wrapped protein kinase C (protein kinase C) into p...

AI Technical Summary

Problems solved by technology

This method is not suitable for collecting a large number of polyhedrons from insect cells. Since many cells are only infected with one virus during mixed infection, many empty polyhedrons will be formed, so that the encapsulation efficiency is low.

Domestic scholars mainly use the silkworm Bacmid expression system to wrap the target protein with the polyhedron protein of the silkworm itself. This method produces a large amount of polyhedron and can be produced by the silkworm itself, but the target protein wrapped with the polyhedron of the silkworm itself is not exclusive. First, it encapsulates a large number of silkworm virus particles, which is not suitable for the current international trend of exclusively encapsulating target proteins through polyhedrons, nor can it be applied to the encapsulation requirements of modern pharmaceutical proteins or vaccine antigens

[0004] The encapsulation process of the target protein by the polyhedron in the cell requires the assistance of a signal peptide protein. The current technical means basically use a 42-93 amino acid or H1 helix at the N-terminus of the target protein as a signal. Peptides, due to the relatively large structure of these proteins, may affect the correct folding process of many target proteins, resulting in the loss of biological activity of the target protein

[0005] Most of the biologically active proteins, such as: GM-CSF, interferon, interleukin, subunit vaccines, etc., are currently mainly administered by injection, which is inconvenient for treatment and difficult to administer for a long time.

We have not seen any research reports on the use of insects to produce polyhedron-encapsulated drugs

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