Chimeric Polypeptide YxxL Motif Exosome Targeting
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Solution Overview
Problem
Current methods lack an effective tool to target and deliver antigenic proteins to exosomes for immunization, particularly in cells like HEK293, limiting the production of immunogenic exosomes for vaccination and immune response induction.
Innovation Solution
A chimeric polypeptide comprising a peptide or protein of interest, a membrane domain for anchoring in the lipid bilayer, and a cytoplasmic domain with a YxxL motif, enabling efficient secretion with exosomes in various eukaryotic cells, including HEK293, by targeting the exosome-forming vesicles.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods are used to deliver antigenic proteins to exosomes, then the production of immunogenic exosomes is limited, but the complexity of the delivery system remains low
Solution Approach 1:
The delivery system is segmented into three functional domains within the chimeric polypeptide: an antigenic protein domain, a transmembrane anchoring domain, and a cytoplasmic sorting domain containing YxxL motifs. This segmentation allows each domain to perform its specific function independently, enabling efficient targeting and delivery of antigenic proteins to exosomes without requiring complex external delivery systems
Solution Approach 2:
The chimeric polypeptide acts as an intermediary molecule that bridges the antigenic protein and the exosome membrane. The transmembrane domain anchors the chimera in the membrane, while the cytoplasmic domain with YxxL motifs mediates sorting into exosomes, thereby facilitating the delivery of antigenic proteins to exosomes through a single integrated molecule rather than complex multi-component systems
2Productivity
If chimeric polypeptide with YxxL motif is used, then secretion with exosomes is significantly amplified, but the polypeptide structure becomes more complex
Solution Approach 1:
The chimeric polypeptide applies local quality by concentrating the sorting function in a specific local region - the cytoplasmic domain containing YxxL motifs. This localized functional element is sufficient to direct the entire chimera into exosomes, achieving high secretion efficiency without requiring complexity throughout the entire polypeptide structure
Solution Approach 2:
The invention changes the parameter of polypeptide structure by introducing specific amino acid sequences (YxxL motifs) that alter the sorting behavior of the protein. This parameter change in the cytoplasmic domain transforms the secretion efficiency from low to high, demonstrating how specific sequence modifications can dramatically enhance exosome association without requiring overall structural complexity
Data Source
AI summary
The present invention concerns a chimeric polypeptide comprising several polypeptide domains, which is capable of being secreted in association with membrane vesicles and in particular exosomes.The present invention also concerns a polypeptide constituted by one of said domains for the secretion of a peptide or polypeptide of interest in association with membrane vesicles and in particular exosomes.The invention also concerns the use of polypeptides of the invention and of polynucleotides coding for said polypeptides for the production of immunogenic compositions based on exosomes or based on DNA or for screening protein interactions.The present invention also concerns exploiting the properties of exosomes comprising a polypeptide of the invention and of immunogenic compositions of the invention for the prophylaxis and/or treatment of an infection by a pathogenic agent, a pathogenic organism, a tumour antigen or a cytoplasmic antigen, in particular to elicit or promote, in vivo, in a host, in particular in a human or non-human mammal or a bird, a humoral and/or cellular response against a virus, bacterium, parasite or tumour.