If the bending of the ssDNA is important for substrate recognition, dependence of binding affinity on substrate length may be expected. To determine if the DNA beyond the four-nucleotide signature sequence contributed to the binding, the length of the ssDNA that contained the recognition sequence was varied in Poly A-TTC. A competition assay with different length oligonucleotides was performed to test the effect of ssDNA length on affinity for substrate (Supplementary Fig. 3). Length was varied from 1 nucleotide flanking each end of TTCA (TTCAA and ATTCA) to 3 nucleotides flanking each end, increasing by one nucleotide addition on either end. Surprisingly, a single nucleotide flanking TTCA signature sequence was not enough to permit binding (Supplementary Fig. 3a), and even three nucleotides on either side still did not bring A3A binding to original binding affinity as Poly A-TTC (AAA TTCA AAA AAA) (Supplementary Fig. 3b). Thus, binding affinity is impacted beyond the recognition motif to prefer longer sequences, although the additional nucleotides are not expected to have any direct contacts with A3A, consistent with the model that intra-DNA interactions modulate A3A affinity.