Fig. 4: Differential contact maps and functional analysis reveals sequence-adaptive DNA binding by AflR-DBD.

a Differential residue contact maps highlight differences in interaction patterns between AflR-DBD binding to vbs vs. ver1 and norA vs. ver1 promoter regions. The y-axis represents residues from AflR-DBD molecules bound at site A and site B. The x-axis labels C and D denote sense and antisense DNA strands. Red indicates higher contact probability in vbs or norA, while blue represents higher contact probability in ver1. Cyan arrows highlight key differential contacts in the terminal regions. b DNA truncation analysis quantifying the relative importance of 5′ vs. 3′ contacts across promoters. Schematic representation of 3′-deletion (3′-Del) and 5′-deletion (5′-Del) constructs are shown with conserved CG sites highlighted in color. EMSA results demonstrate differential effects of these truncations across the three promoters. The ratio of stable complex band intensity (arrows) for 3′-deletion vs. 5′-deletion constructs is quantified in the graph. EMSA experiments were performed independently in triplicate, with one representative image shown and quantitative ratios from all three experiments plotted. c Mutational analysis identifies key residues required for DNA recognition. Binding curves from EMSA titration experiments show differential effects of mutations in the N-terminal (R23A, D27A), zinc cluster (K36A, V37A), and C-terminal regions (R63A, G65A, R66A) across all three promoters. Error bars represent the Mean ± SEM from three independent experiments.