On Relevance of Codon Usage to Expression of Synthetic and Natural Genes in Escherichia coli

A recent investigation concluded that codon bias did not affect expression of green fluorescent protein (GFP) variants in Escherichia coli, while stability of an mRNA secondary structure near the 5' end played a dominant role. We demonstrate that combining the two variables using regression trees or support vector regression yields a biologically plausible model with better support in the GFP data set and in other experimental data: codon usage is relevant for protein levels if the 5' mRNA structures are not strong. Natural E. coli genes had weaker 5' mRNA structures than the examined set of GFP variants and did not exhibit a correlation between the folding free energy of 5' mRNA structures and protein expression.