Complementary codes vs. amino acid co-locations. (A) The propensity of the 400 possible amino acid pairs was monitored in 80 different protein structures with the SeqX tool. The tool detected co-locations when two amino acids were closer than 6 Å to each other (neighbors on the same strand were excluded). The total number of co-locations was 34,630. Eight different complementary codes were constructed for the codons (two optimal and six suboptimal). In the two optimal codes all three codon residues (123) were complementary (C) or reverse-complementary (RC) to each other. In the suboptimal codes only two of three codon residues were C or RC to each other (12, 13, 23), while the third was not necessarily complementary (X). (For example, complementary code RC_1X3 means that the first and third codon letters are always complementary, but not the second, and the possible codons are read in reverse orientation). The 400 co-locations were divided into 20 subgroups corresponding to 20 amino acids (one of the co-locating pairs), each group containing the 20 amino acids (corresponding to the other amino acid in the co-locating pair). If the codons of the amino acid pairs followed the predefined complementary code, the co-location was regarded as positive (P); if not, the co-location was regarded as negative (N). Each symbol represents the mean frequency of P or N co-locations corresponding to the indicated amino acid. Paired Student's t-test, n = 20 (see Fig. 2 for explanation). (B) The ratio of positive (P) and negative (N) co-locations was calculated on data from (A). Each bar represents the mean ± SEM, n = 20.