E. coli Strain Engineering for Recombinant Protein Secretion
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Solution Overview
Problem
Current methods for producing recombinant proteins in E. coli face challenges with contamination from host proteins, which complicates purification and reduces yield, as these proteins compete for secretion apparatus components and are often immunogenic or toxic.
Innovation Solution
Development of E. coli strains with reduced expression of host proteins like YddS, achieved through genetic modifications such as deletion or mutation, to minimize their secretion and contamination in the recombinant protein product, thereby enhancing the specific yield and purity of recombinant proteins.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If host proteins are naturally secreted by the bacterium, then the secretion apparatus is utilized, but the recombinant protein secretion is reduced due to competition for secretion components
Solution Approach 1:
The patent removes specific host protein genes (oppA, dppA, yddS) from the E. coli genome to eliminate competing proteins that utilize the secretion apparatus. This extraction of problematic elements directly increases available secretion capacity for the recombinant protein while reducing contamination in the final product.
Solution Approach 2:
The patent modifies the genetic composition of the host cell by deleting specific genes, thereby changing the parameter of host protein secretion levels. This genetic modification alters the secretion landscape to favor recombinant protein production without compromising cellular viability.
2Manufacturing precision
If multiple purification stages are used to remove host proteins, then the purity of recombinant protein increases, but the production cost and process complexity increase
Solution Approach 1:
The patent performs preliminary action by genetically modifying the host cell before protein production to prevent the secretion of contaminating host proteins. This preemptive measure reduces the burden on downstream purification processes, allowing for simpler and more cost-effective purification protocols.
3Manufacturing precision
If host proteins are deleted to reduce contamination, then the purity of recombinant protein increases, but the host cell's physiological functions may be compromised
Solution Approach 1:
The patent applies local quality by selectively deleting only specific host protein genes (oppA, dppA, yddS) that are responsible for contamination, while preserving the rest of the host cell's physiological functions. This targeted approach maintains cell viability and metabolic functionality while achieving the desired purification effect.
Data Source
AI summary
The invention relates to a strain of microorganism comprising a gene encoding a recombinant protein and a mutated gene encoding a host protein which is not a protease, characterized in that the recombinant protein is secreted in a fermentation and the mutated gene encoding the host protein is mutated in such a way that it causes a reduced expression of the host protein compared to the wild type gene underlying the mutated gene.