Proteins play a crucial role in cell signaling and inflammation reactions. Today, recombinant proteins are applied in all biological and biomedical science. Since proteins are different, you must optimize the purification strategies for every protein depending on the intended use. However, this process can be a challenge, demanding enormous investments in time and costs.
Here are the tips for optimizing protein production during recombinant protein expression.
- Add stability sequences
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The ultimate stage of protein production is to improve protein purification service in a well-buffered solution. You can use several tags to enhance protein solubility and purification. The primary aim is to enhance the protein solubility that will enhance the expression. For example, Protein A with 280 amino acids can reinforce the solubility of a protein without interfering with its folding. Peptide sequencing can also be suitable in overcoming the problems related to protein solubility. It amplifies the polymerization and solubility characteristics of the amino acids.
- Change expression conditions
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Using lower expression temperatures can affect the production by limiting protein aggregation and synthesis rate. Therefore, focus on using higher expression temperatures to improve protein production. High cultivation temperatures can increase protein expression by increasing replication, transcription, and translation rates. It also plays a crucial role in protein production yields by improving bacteria growth.
Therefore, focus on using higher inducible promoters that maximize protection under high-temperature conditions. Under high expression levels, the protein will accumulate in inclusion bodies for better yield.
- Optimize the codons
Codon optimization entails switching those used in a transgene without affecting the amino acid sequence. It will increase the protein abundance in the host organism by replacing the rare with abundant codons. During recombinant protein expression, these codons encode most of the amino acids. Hence, you will have multiple tRNAs corresponding to an amino acid. The idea is to optimize codon usage of the gene and enhance its translation rate. It will exploit the tRNA content ratio and increase the mRNA elongation speed in the expression system.
- Choose the correct expression vector.
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You must use the proper expression factor during protein production and expression. With many plasmids available on the market, select the right one from promoters, replicons, and selection markers. For example, the lac promoter is crucial in lac operon to drive protein expression.
Besides, you can also use the T7 promoter that is hugely popular for recombinant protein expression. Using strong expression promoters and high inducer concentrations can increase protein concentrations. This is crucial in enhancing protein production.
- Chose an adaptable bacterial strain
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One way of enhancing the quality of proteins in E. coli is to select the best bacteria strains. There are several strategies developed with plenty of bacteria strains. Each variant has characteristics that facilitate the production of peptides with rare codons. Also, focus on producing the proteins within a specific condition that offers the best bacterial growth and protein expression.
The Bottom Line
It is easy to optimize protein production in E. coli if you have the necessary strategies. The idea is to provide the best optimal conditions to recombine protein expression. Since each protein requires its optimal condition, find the right expression conditions for the expression, production, and purification. These tips can help you optimize production with ease.