Structural and Binding Properties of Dps - A Nucleoid Associated Protein

Area of Science:

• Microbiology
• Structural Biology
• Biophysics

Background:

• Prokaryotic DNA compaction relies on nucleoid-associated proteins (NAPs).
• DNA-binding proteins from starved cells (Dps) are key NAPs with ferroxidase activity, forming dodecameric assemblies to protect the genome under stress.
• Understanding Dps function requires analyzing its structural and binding dynamics under varying environmental conditions.

Purpose of the Study:

• To investigate the structural and biophysical properties of *Escherichia coli* Dps across a wide pH range (3-11).
• To elucidate how pH affects Dps assembly, DNA binding affinity, and genome compaction mechanisms.
• To explore the role of Dps as a pH-responsive factor in bacterial nucleoid organization.

Main Methods:

• Single-particle cryo-electron microscopy (cryo-EM) for high-resolution structural analysis (~1.75 Å).
• Adaptive Poisson-Boltzmann electrostatic modeling to predict surface charge distribution.
• Electrophoretic mobility shift assays (EMSAs) to quantify DNA binding affinity.
• Cryo-electron tomography (cryo-ET) to visualize Dps-DNA complex formation.

Main Results:

• The canonical ferritin-like fold of Dps remains stable across the tested pH range (3-11).
• Surface electrostatics shift from positive to negative with increasing pH, significantly modulating DNA binding affinity (EC50 values range from 73.4 nM at pH 5 to 815.5 nM at pH 11).
• Dps forms amorphous ~50 nm globular or tubular complexes with short DNA fragments, contrasting with lattice co-crystals seen with longer DNA scaffolds.

Conclusions:

• Dps functions as a pH-sensitive regulator of bacterial nucleoid compaction.
• A two-stage assembly model is proposed: protonation-driven nucleation at N-terminal lysines, followed by lattice ordering on extended DNA.
• These findings highlight the dynamic nature of Dps in adapting genome organization to environmental pH fluctuations.