Structural and Binding Properties of Dps: A Nucleoid Associated Protein

Prokaryotic chromosomal DNA is compacted and protected by nucleoid-associated proteins (NAPs). Among these, the DNA-binding proteins from starved cells (Dps) combine ferroxidase activity with nonspecific DNA binding to form dodecameric assemblies that condense and safeguard the genome under stress. Here, we present an integrated structural and biophysical analysis of Escherichia coli (E. coli) Dps across a pH range from 3 to 11, combining (i) single-particle cryo-EM reconstructions at [~]1.75 [A] resolution, (ii) adaptive Poisson-Boltzmann electrostatic modelling, (iii) electrophoretic mobility shift assays (EMSAs) on a 448 bp DNA fragment, and (iv) cryo-electron tomography (cryo-ET) of Dps-DNA assemblies. Through this analysis, we show that (1) the canonical ferritin-like fold is maintained at all pH values; (2) surface electrostatics shift from highly positive to net negative as the pH increases, modulating DNA affinity (EC50 values of 73.4 nM at the lowest measurement of pH 5, to 815.5 nM at the highest measurement of pH 11); and (3) short-fragment Dps-DNA complexes form amorphous [~]50 nm globular or tubular complexes rather than lattice co-crystals observed on kilobase-length DNA scaffolds. The findings reveal a possible two-stage assembly model, in which protonation-driven nucleation occurs via the Dps N-terminal lysine residues, followed by lattice ordering on long DNA. Our results further define Dps as a pH-responsive nucleoid compaction factor in bacteria.