*2.4. Comparison of Frequency of Binding in the DOT Region and Other Residues of a Protein*

To estimate the difference between binding in DOT regions and other part of proteins, we calculated the binding frequency of amino acids in these regions, as shown in Figure 6 and Figure S5. Amino acids significantly differ in their binding with RNA in DOT regions and in the complete protein (*p*-value for the mean is less than 0.01). In non-ribosomal proteins, when the 3.5 Å cut-off is considered, nonpolar and aromatic residues mostly have high frequency values in the DOT regions than in the overall protein. All the frequencies are observed to be significant when statistical analysis is performed for the bootstrapped sample of the frequencies (*p*-value is less than 0.01). Residues such as His, Phe, and Leu are found to have a more than 3-fold increase in the frequency of binding in the DOT regions than in other parts of the proteins. A similar trend is observed in the NR6 dataset (Figure S5).

**Figure 6.** Binding frequency of each amino acid in the DOT region and in the overall protein for non-ribosomal complexes using the 3.5 Å cut-off.
