Codon optimization can improve expression of human genes in Escherichia coli: A multi-gene study

Area of Science:

• Molecular Biology
• Biotechnology
• Protein Expression

Background:

• Biased codon usage in Escherichia coli (E. coli) can limit heterologous protein production efficiency.
• Traditional methods to address rare codons include gene mutagenesis or introducing specific transfer RNAs (tRNAs).
• Advances in synthetic gene technology offer a cost-effective alternative for optimizing gene expression.

Purpose of the Study:

• To compare the expression efficiency of native versus codon-optimized synthetic human short-chain dehydrogenase/reductase (SDR) genes in E. coli.
• To evaluate the impact of rare codon tRNAs on the expression of native genes.
• To determine if synthetic gene optimization is superior to using rare codon tRNAs for enhancing protein production.

Main Methods:

• Analyzed 30 human SDR genes, comparing native and synthetic (codon-optimized) versions.
• Constructed expression vectors with N-terminal polyhistidine tags for purification.
• Compared protein expression in standard E. coli and E. coli strains supplemented with a rare codon tRNA plasmid (pRARE2).

Main Results:

• Synthetic genes generally showed improved expression and easier purification in standard E. coli compared to native genes.
• Expression of native genes with added rare codon tRNAs often mimicked the performance of synthetic genes.
• Native gene expression was superior in only one out of the 30 SDR genes studied.

Conclusions:

• Altering codon preference through synthetic gene design can enhance heterologous protein expression in bacteria.
• The benefits of codon optimization can often be replicated by supplementing the host E. coli with rare codon tRNAs.
• Both strategies offer viable approaches to improve the production of challenging heterologous proteins.