Optimization of translation profiles enhances protein expression and solubility

Area of Science:

• Molecular Biology
• Biochemistry
• Protein Engineering

Background:

• Messenger RNA (mRNA) translation speed is known to be non-uniform.
• This non-uniformity is increasingly recognized for its role in coordinating co-translational protein folding.
• Understanding this process is crucial for optimizing protein expression in biotechnology.

Purpose of the Study:

• To investigate the impact of mRNA translation speed on protein folding and solubility.
• To explore methods for enhancing protein solubility and yield through codon optimization.
• To demonstrate that mRNA contains information beyond the amino acid sequence for protein expression.

Main Methods:

• Structure-based homology modeling to identify protein domains in epoxide hydrolases (EHs).
• Introduction of slow-translating codons to control domain translation speed.
• Synonymous codon substitutions to alter translation speed.
• Analysis of protein solubility and yield in Escherichia coli.

Main Results:

• Engineered slow-translating codons significantly improved the solubility of two metagenomic EHs in E. coli.
• Altering translation speed in an EH from Agrobacterium radiobacter demonstrated the importance of transient attenuation for solubility.
• Converting slow-translating regions to fast-translating codons resulted in insoluble protein.
• Decreasing the free folding energy of the 5'-coding region enhanced ribosomal loading and protein yield.

Conclusions:

• mRNA translation speed is a critical factor coordinating co-translational folding, influencing protein solubility and expression levels.
• Codon optimization by modulating translation speed offers a powerful strategy for enhancing recombinant protein production.
• This study reveals a new layer of information encoded within mRNA that governs protein biogenesis.