Bacillus Fermentation Temperature Shift for Protein Yield
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Solution Overview
Problem
Current large-scale industrial fermentation processes for producing proteins of interest using Bacillus host cells face challenges in maximizing yield, particularly in maintaining optimal growth and expression conditions to minimize inclusion body formation and enhance protein production.
Innovation Solution
A method involving a two-phase cultivation process where Bacillus host cells are inoculated into a fermentation medium with an expression construct, with the first phase conducted at a first temperature and the second phase at a higher temperature, utilizing feed solutions to manage carbon source delivery and promote continuous protein expression.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If constant temperature cultivation is used, then process simplicity is maintained, but protein yield is limited
Solution Approach 1:
The patent applies dynamic temperature control by shifting from a first temperature (e.g., 30°C) during the first cultivation phase to a second temperature (e.g., 37°C) during the second cultivation phase. This dynamic adjustment optimizes protein yield by matching temperature conditions to different growth and expression stages, resolving the contradiction between maintaining process simplicity and achieving high productivity.
2Reliability
If temperature is decreased, then inclusion body formation is reduced, but protein expression efficiency decreases
Solution Approach 1:
The patent segments the cultivation process into two distinct phases with different temperature conditions. The first phase uses a lower temperature to promote soluble protein expression and reduce inclusion body formation, while the second phase uses a higher temperature to enhance overall protein expression rate. This temporal segmentation allows both contradictory requirements to be satisfied at different stages.
Solution Approach 2:
The patent implements periodic temperature adjustment, switching between two temperature regimes at a defined transition point during cultivation. This periodic action enables the system to alternate between conditions optimized for soluble expression and conditions optimized for high expression rate, achieving both reliability and productivity.
3Productivity
If feed solution addition is increased, then carbon source availability is improved, but process control complexity increases
Solution Approach 1:
The patent employs fed-batch cultivation with pre-planned feed solution addition strategies. Carbon source is added in controlled amounts at predetermined time points or based on predefined criteria (e.g., glucose concentration thresholds). This preliminary planning of feed addition reduces real-time control complexity while maintaining high productivity through optimized carbon source availability.
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
This approach significantly increases protein yield, with temperature shifts enhancing production by up to 400% compared to constant temperature processes, improving the efficiency of protein production in industrial fermentation.
Implementation Method 1
The biotechnological production of the product of interest is conducted via fermentation and subsequent purification of the product. Microorganisms, like the Bacillus species, are capable of secreting significant amounts of product into the fermentation broth.
Data Source
AI summary
The present invention relates to the field of industrial fermentation. In particular, it relates to a method for cultivating a Bacillus host cell comprising the steps of (a) inoculating a fermentation medium with a Bacillus host cell comprising an expression construct for a gene encoding a protein of interest, (b) cultivating for a first cultivation phase the Bacillus host cell in said fermentation medium under conditions conducive for the growth of the Bacillus host cell and the expression of the protein of interest, wherein the cultivation of the Bacillus host cell comprises the addition of at least one feed solution and wherein the cultivation during the first cultivation phase is carried out at a first temperature, and (c) cultivating for a second cultivation phase the Bacillus host cell culture obtained in step (b) under conditions conducive for the growth of the Bacillus host cell and the expression of the protein of interest, wherein the cultivation comprises the addition of at least one feed solution and wherein the cultivation during the second cultivation phase is carried out at a second temperature, said second temperature being higher than the first temperature. The invention also provides for a Bacillus host cell culture obtainable by the said method.