Recombinant Bacterium Regulated Antigen Expression Colonization
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Recombinant attenuated bacterial vaccines face challenges in effectively colonizing lymphoid tissues due to susceptibility to environmental stresses and overexpression of antigens, leading to reduced immunogenicity and plasmid loss, which diminishes the immune response.
Innovation Solution
A recombinant bacterium with chromosomally integrated nucleic acid sequences encoding a repressor and a vector with an antigen operably linked to a regulatable promoter, allowing controlled expression and attenuation to optimize antigen production and colonization of lymphoid tissues.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If the recombinant nucleic acid sequence is highly expressed to produce high levels of antigen, then the immunogenicity is improved, but the microorganism becomes susceptible to environmental stresses and shows deleterious effects
Solution Approach 1:
The patent applies dynamic regulation of gene expression by using inducible promoters (such as lac, trc, tac, or araBAD promoters) that allow the recombinant nucleic acid sequence to be expressed at high levels only when induced, while remaining repressed under normal growth conditions. This dynamic control enables the microorganism to maintain high antigen production capability without suffering the deleterious effects of constitutive high-level expression, thereby resolving the contradiction between antigen quantity and microorganism reliability
Solution Approach 2:
The patent changes the expression parameter from constitutive to regulated/inducible by incorporating control elements (promoters, operators, repressors) that respond to environmental signals. This parameter change allows the system to adjust antigen production levels based on growth conditions, maintaining both high immunogenicity and microorganism viability
2Object-affected harmful factors
If the recombinant microorganism is attenuated to reduce pathogenicity, then the safety is improved, but the ability to colonize lymphoid tissues is reduced
Solution Approach 1:
The patent employs dynamic regulation of attenuation genes through inducible promoters, allowing the microorganism to exhibit different phenotypes in different environments. Under in vitro conditions, the attenuation genes are repressed, maintaining full colon化 capability. Upon induction or in host conditions, the attenuation genes are expressed, reducing pathogenicity. This dynamic switching resolves the contradiction between safety and colonization ability
Solution Approach 2:
The patent implements periodic expression of attenuation genes during the infection cycle, where the genes are repressed during initial colonization phase to maintain productivity, then induced later to provide attenuation and safety. This temporal separation of functions allows the microorganism to first colonize effectively, then become attenuated, resolving the contradiction
3Quantity of substance
If plasmid vectors are used to increase antigen expression, then the immunogenicity is improved, but plasmid loss occurs leading to reduced immune response
Solution Approach 1:
The patent extracts the antigen-encoding nucleic acid sequence from plasmid vectors and integrates it directly into the bacterial chromosome. This extraction eliminates the plasmid component that causes instability and loss, while maintaining high antigen expression levels through chromosomal integration and regulated promoters. The solution resolves the contradiction by removing the source of plasmid loss while preserving high antigen quantity
Solution Approach 2:
The patent merges the antigen-encoding sequence with the bacterial chromosome through integration, creating a stable heritable unit. This merging eliminates the separation between plasmid and chromosomal DNA, ensuring stable inheritance and retention of the antigen gene across generations, thereby resolving the plasmid loss issue while maintaining high immunogenicity
Data Source
AI summary
The invention relates to compositions and methods for making and using recombinant bacteria that are capable of regulated attenuation and/or regulated expression of one or more antigens of interest.