PrsA Lipoprotein Anchoring for Stable Display on Lactic Acid Bacteria
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Solution Overview
Problem
Existing technologies face challenges in stably and efficiently displaying target proteins in large quantities on the cell surface of microorganisms, particularly lactic acid bacteria, limiting their applications in vaccine development and biocatalysis due to safety concerns and insufficient immune responses.
Innovation Solution
A recombinant vector system utilizing the PrsA promoter and PrsA protein from lactic acid bacteria, which are GRAS microorganisms, to anchor and display target proteins on the cell surface, enabling stable and high-expression levels of target proteins.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If transmembrane anchor proteins are used to display target proteins on the cell membrane, then the target proteins can be anchored to the membrane, but the membrane rigidity is significantly affected and large-quantity display is limited
Solution Approach 1:
The patent introduces a lipoprotein anchor system as an intermediary between the target protein and the cell membrane. This lipoprotein anchor mediates the attachment by forming a covalent bond between the target protein and lipid molecules in the membrane, rather than directly inserting into the membrane like transmembrane anchors. This intermediary approach allows large-quantity display without compromising membrane rigidity.
Solution Approach 2:
The patent extracts the anchoring function from the transmembrane domain and separates it into a distinct lipoprotein anchor component. By taking out the hydrophobic transmembrane insertion requirement and replacing it with a lipid-covalent bond mechanism, the system enables high-density display while preserving membrane properties.
2Productivity
If conventional expression systems are used to increase target protein expression, then expression levels can be improved, but stable display on cell surface in large quantities remains difficult
Solution Approach 1:
The patent changes the anchoring mechanism parameter from transmembrane insertion to lipoprotein covalent bonding. This parameter change fundamentally alters how proteins are attached to the membrane, enabling both high expression levels and stable cell surface display by using the cysteine residue-covalent bond-lipid mechanism rather than relying solely on secretion signals and membrane insertion efficiency.
3Object-affected harmful factors
If lactic acid bacteria are used as host organisms, then safety concerns are minimized due to GRAS status, but the ability to display target proteins in large quantities on cell surface is insufficient
Solution Approach 1:
The patent creates a universal cell surface display system using lactic acid bacteria that can display various target proteins in large quantities. The lipoprotein anchor system developed herein provides multi-functionality, enabling these safe GRAS organisms to serve both as safe delivery vehicles and as high-capacity display platforms for vaccines and therapeutics, overcoming the previous limitation of low display capacity.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The system allows for stable and high-level display of target proteins on lactic acid bacteria, enhancing their utility in pharmaceutical applications such as therapeutic agents, immunotherapeutics, and vaccines, while minimizing safety risks.
Implementation Method 1
a PrsA promoter that induces overexpression of PrsA
Implementation Method 2
the -SH group of a cysteine residue present immediately behind a secretion signal is covalently bonded to the carbon of the glycerin head of a lipid
Data Source
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AI summary
The present invention relates to: lactic acid bacteria extracellular membrane protein PrsA having a cell surface display anchor motif function; and a vector system for cell surface display of a target protein, the system comprising a sequence that encodes the PrsA and a strong promoter that is upstream of the PrsA gene.