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pCODE tRNA Expression Plasmids Compensate for Rare Codons in Recombinant Protein Production.

Marilena Feller1, Ana G V Sepulchro1, Maurice Mürköster2

  • 1BRIGHT, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.

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|April 21, 2026
PubMed
Summary
This summary is machine-generated.

This study introduces pCODE plasmids for tRNA overexpression in Escherichia coli, effectively complementing rare codons to enhance recombinant protein production with minimal negative effects. This offers a new tool for molecular biology and genetic regulation.

Keywords:
Escherichia coliGFPcodon optimizationrecombinant protein productiontRNA expressiontranscription

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • Synonymous codon usage significantly impacts recombinant protein production.
  • Rare codons are often avoided in synthetic gene design, but alternative strategies like tRNA overexpression are less understood.
  • Understanding codon usage is crucial for optimizing protein expression.

Purpose of the Study:

  • To develop and characterize a versatile plasmid collection (pCODE) for tRNA overexpression in Escherichia coli.
  • To investigate the consequences and limitations of tRNA overexpression for compensating suboptimal codon usage.
  • To explore the potential of inducible tRNA expression for genetic regulation.

Main Methods:

  • Development and characterization of pCODE plasmids for tRNA gene overexpression in Escherichia coli.
  • Testing the efficacy of pCODE plasmids in complementing rare codons across various gene constructs.
  • Analysis of limitations and negative effects associated with tRNA overexpression.

Main Results:

  • pCODE plasmids effectively complement rare codons with minimal limitations, irrespective of rare codon characteristics.
  • The position, type, number, and distribution of rare codons did not significantly hinder complementation.
  • One pCODE plasmid demonstrated inducible tRNA expression, suggesting a novel regulatory mechanism.

Conclusions:

  • pCODE plasmids are a versatile tool for enhancing recombinant protein production by addressing suboptimal codon usage.
  • tRNA overexpression is a viable strategy to improve protein yields without significant drawbacks.
  • Inducible tRNA expression via pCODE plasmids offers a new avenue for genetic regulation.