Shikimate Pathway Engineering for Aromatic Amino Acid Yield
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Solution Overview
Problem
Current methods for improving microbial production of aromatic amino acids, such as L-tyrosine, are inefficient due to the complexity of the shikimate pathway, which is difficult to engineer and often results in the introduction of new bottlenecks, limiting yield optimization.
Innovation Solution
Engineering microorganisms with recombinant polynucleotides containing operons that encode enzymes for the shikimate biosynthesis pathway, optimizing gene expression and plasmid copy number, and modifying enzymes like YdiB and AroB to relieve bottlenecks, allowing for increased production of shikimate and aromatic amino acids.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional step-wise metabolic engineering is used to improve aromatic amino acid production, then some yield improvement can be achieved, but the process is time-consuming and leads to local yield maxima rather than global optimal yield
Solution Approach 1:
The patent applies preliminary action by using computational algorithms to predict and identify multiple potential bottleneck genes before experimental validation. The system pre-calculates the effects of deleting or overexpressing genes based on metabolic flux analysis, allowing researchers to prioritize which genes to modify first, thereby reducing the time-consuming trial-and-error process of traditional step-wise engineering.
Solution Approach 2:
The patent implements feedback through an iterative computational-experimental cycle. The algorithm predicts bottleneck genes based on current metabolic flux data, experiments validate these predictions by measuring actual product yields, and the results feed back into the model to refine flux predictions. This closed-loop feedback system enables the methodology to escape local optima and converge toward global optimal yield by continuously adjusting gene modification strategies based on experimental outcomes.
2Productivity
If multiple gene modifications are made to overcome bottlenecks in the shikimate pathway, then production yield can be improved, but new bottlenecks are introduced and the pathway becomes difficult to engineer
Solution Approach 1:
The patent applies segmentation by dividing the complex shikimate pathway into discrete functional segments or modules, each with identifiable bottleneck genes. The computational algorithm analyzes flux distribution across these segments and identifies specific genes within each segment that limit overall pathway performance. This modular segmentation allows researchers to tackle pathway optimization in manageable units rather than attempting to modify the entire pathway simultaneously, reducing engineering complexity.
Solution Approach 2:
The patent employs parameter changes by using computational algorithms to predict how modifying specific gene parameters (expression levels, enzyme activity) will affect overall pathway flux. The system calculates optimal parameter values for multiple genes simultaneously, considering their interdependent effects on metabolic flux. This allows for coordinated modification of multiple genes with predicted synergistic effects, improving pathway flux while managing complexity through rational parameter optimization rather than random modifications.
Data Source
AI summary
The present disclosure relates to engineered microorganisms that produce amino acids and amino acid intermediates. In particular, the disclosure relates to recombinant nucleic acids encoding operons that increase production of aromatic amino acids and the aromatic amino acid intermediate shikimate; microorganisms with increased production of aromatic amino acids and the aromatic amino acid intermediate shikimate; and methods related to the production of aromatic amino acids, the aromatic amino acid intermediate shikimate, and commodity chemicals derived therefrom.