To gain further insight into the functional effects of TFBS mutations, we used a dataset that mapped human CTCF binding sites across four individuals from [16] (see Materials and methods for more details). TFBS mutations detected in this dataset often did not result in a significant loss of binding, with approximately 75% of mutated sites retaining at least two-thirds of the binding signal. This was particularly prominent at conserved sites (BLS >0.5), 90% of which showed this 'buffering' effect (Figure 5a). To address whether buffering could be explained solely by the flexibility of CTCF sequence preferences, we analyzed between-allele differences in the PWM score at polymorphic binding sites. As expected, globally CTCF binding signal correlated with the PWM score of the underlying motifs (Figure S6A in Additional file 1). Consistent with this, alleles with minor differences in PWM match generally had little effect on the binding signal compared to sites with larger PWM score changes (Figure 5b), suggesting that the PWM model adequately describes the functional constraints of CTCF binding sites. At the same time, we found that CTCF binding
