Plant VLP Production via M2 Cysteine-to-Serine Mutation
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Solution Overview
Problem
Current methods for producing viral proteins, such as influenza virus-like particles (VLPs), face challenges in achieving high yields and effective immunogenicity due to issues like incorrect protein folding and reduced antigen presentation, particularly when co-expressing M2 protein, which can inhibit HA accumulation and VLP production.
Innovation Solution
A method involving the co-expression of a structural virus protein, like HA, with a channel protein (e.g., M2 or BM2) in plants, utilizing specific regulatory regions and enhancers to increase VLP production and stability, and incorporating plant-specific N-glycans for enhanced immunogenicity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If M2 protein is co-expressed with HA to form VLPs, then VLP structure and immunogenicity are improved, but HA accumulation and VLP production are reduced
Solution Approach 1:
The patent modifies the M2 protein sequence by substituting cysteine residues at positions 17 and 19 with serine, creating M2(S17S19). This parameter change in the protein structure eliminates the harmful interaction between M2 and HA while preserving the ion channel function and VLP formation capability, thereby resolving the contradiction between VLP structure integrity and HA accumulation
Solution Approach 2:
The patent converts the harmful overexpression effect of M2 (which blocks HA transport and reduces VLP production) into a beneficial outcome by using the mutated M2(S17S19) variant. The mutation eliminates the toxic interaction while maintaining the essential functions of M2 in VLP assembly and immunogenicity, thus transforming a harmful factor into a beneficial one
2Reliability
If M2 protein is overexpressed to enhance VLP formation, then ion channel function is improved, but intracellular transport of HA is inhibited
Solution Approach 1:
The patent changes the amino acid sequence parameters of M2 by substituting cysteine residues with serine at positions 17 and 19. This parameter modification maintains the ion channel function (as evidenced by pH gradient formation capability) while eliminating the inhibition of HA intracellular transport, allowing both functions to coexist without interference
3Productivity
If plant system is used for VLP production, then scalability and cost are improved, but protein folding and antigen presentation are reduced
Solution Approach 1:
The patent modifies the M2 protein sequence parameters (cysteine to serine substitution) to adapt it for optimal function in the plant expression system. This parameter change ensures proper protein folding and interaction with HA in plants, while maintaining the scalability and cost advantages of plant-based production
Solution Approach 2:
The mutated M2(S17S19) protein acts as an intermediary that facilitates proper protein folding and assembly of VLPs in the plant system. It mediates between the plant cellular environment and the viral protein components, ensuring correct antigen presentation while maintaining the benefits of plant-based production
Data Source
Figure 1A~1C
Figure 1D
Figure 1E
AI summary
A method of producing a virus like particle (VLP) in a plant is provided. The method comprises introducing a first nucleic acid and a second nucleic acid into the plant, or portion of the plant. The first nucleic acid comprises a first regulatory region active in the plant and operatively linked to a nucleotide sequence encoding a structural virus protein. The second nucleic acid comprises a second regulatory region active in the plant and operatively linked to a nucleotide sequence encoding a channel protein, for example but not limited to a proton channel protein . The plant or portion of the plant is incubated under conditions that permit the expression of the nucleic acids, thereby producing the VLP.