  Background: The secondary structure and complexity of mRNA influences its
accessibility to regulatory molecules (proteins, micro-RNAs), its stability and
its level of expression. The mobile elements of the RNA sequence, the wobble
bases, are expected to regulate the formation of structures encompassing coding
sequences.
  Results: The sequence/folding energy (FE) relationship was studied by
statistical, bioinformatic methods in 90 CDS containing 26,370 codons. I found
that the FE (dG) associated with coding sequences is significant and negative
(407 kcal/1000 bases, mean +/- S.E.M.) indicating that these sequences are able
to form structures. However, the FE has only a small free component, less than
10% of the total. The contribution of the 1st and 3rd codon bases to the FE is
larger than the contribution of the 2nd (central) bases. It is possible to
achieve a ~ 4-fold change in FE by altering the wobble bases in synonymous
codons. The sequence/FE relationship can be described with a simple algorithm,
and the total FE can be predicted solely from the sequence composition of the
nucleic acid. The contributions of different synonymous codons to the FE are
additive and one codon cannot replace another. The accumulated contributions of
synonymous codons of an amino acid to the total folding energy of an mRNA is
strongly correlated to the relative amount of that amino acid in the translated
protein.
  Conclusion: Synonymous codons are not interchangable with regard to their
role in determining the mRNA FE and the relative amounts of amino acids in the
translated protein, even if they are indistinguishable in respect of amino acid
coding.
