Yeast Expression Platform for Full-Length Difficult Proteins
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Solution Overview
Problem
Existing recombinant expression systems struggle to efficiently produce full-length, well-folded, and active forms of difficult to express proteins (DTE-Ps) such as membrane proteins, structural proteins, enzyme proteins, and drug target molecules, leading to aggregation, insoluble expression, and high production costs, with no universal solution for scalable and high-quality production.
Innovation Solution
A versatile yeast-based recombinant expression platform using protease-deficient yeast cells and engineered expression vectors with specific promoters and markers, allowing for the expression of DTE-Ps, including Neuraminidase, VP7, Nav1.7, fatty acid desaturase, and CD59, by integrating or episomal vectors with Gal1 or ADH2 promoters and auxotrophic markers.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional host cells (E. coli, yeast, mammalian, insect) are used for recombinant expression, then well-folded and active proteins can be produced, but difficult to express proteins (with hydrophobicity, high cysteine-proline residues, repetitive amino acids) do not express efficiently
Solution Approach 1:
The invention modifies host cell parameters by creating protease-deficient strains (deleting specific protease genes) and optimizing expression conditions (temperature, induction timing, media composition) to enable efficient expression of difficult to express proteins that conventional systems cannot produce
2Reliability
If varied host systems and methodologies are applied to express difficult to express proteins, then expression can be achieved, but the process becomes unpredictable, laborious, costly and time-consuming
Solution Approach 1:
The invention creates a universal expression platform using protease-deficient yeast cells that can express multiple types of difficult to express proteins (membrane proteins, structural proteins, enzyme proteins, drug targets) through a standardized methodology, eliminating the need for system-specific optimizations
3Reliability
If membrane proteins are expressed as modified proteins with deletion of transmembrane domain in E. coli, then insoluble expression is avoided, but full-length sequence expression is lost and full-length protein for analysis is unavailable
Solution Approach 1:
The invention uses yeast cells as an intermediary host system that naturally handles membrane protein expression better than E. coli, allowing full-length membrane proteins to be expressed in their native form with proper folding and solubility without requiring truncation or modification
4Manufacturing precision
If eukaryotic host systems are used for expression of full-length membrane proteins, then intact and full-length proteins are produced, but scalability and commercial production capability are limited
Solution Approach 1:
The invention uses yeast cells (a simple, fast-growing eukaryotic system) as a disposable production platform that can be rapidly cultured and scaled up, providing both the quality of eukaryotic expression and the scalability needed for commercial production
Data Source
AI summary
The present invention relates to expression of SARS-CoV like virus proteins [S, M and E] proteins; recombinant polynucleotides, polypeptides; constructs, virus-like particles (VLPs); immunogenic compositions or vaccines comprising Virus Like Particles (VLPs). Method of producing the VLPs/expressing the multi-subunit virus like proteins and method for co-expression of multi-subunit and virus like proteins (VLPs) are also provided. The present invention also provides strategies, methods, systems, kits and combinations for scalable expression, purification and enhanced production of the virus like proteins of SARS-CoV while maintaining their size range and composition. Such multi-subunit VLPs can be utilized to make immunogenic compositions or vaccines.