Functional characterization of the DNA-binding protein from starved cells (DPS) as a molecular chaperone under heat stress.

The DNA-binding protein from starved cells, known as DPS, has been characterized as a crucial factor in protecting E. coli from external stresses. The DPS functions in various cellular processes, including protein-DNA binding, ferroxidase activity, compaction of chromosome and regulation of stress resistance gene expression. DPS proteins exist as oligomeric complexes; however, the specific biochemical activity of oligomeric DPS in conferring heat shock tolerance has not been fully understood. Therefore, we investigated the novel functional role of DPS under heat shock. To elucidate the functional role of DPS under heat shock conditions, we purified recombinant GST-DPS protein and demonstrated its thermostability and existence in its highly oligomeric form. Furthermore, we discovered that the hydrophobic region of GST-DPS influenced the formation of oligomers, which exhibited molecular chaperone activity, thereby preventing the aggregation of substrate proteins. Collectively, our findings highlight the novel functional role of DPS, as a molecular chaperone and may confer thermotolerance to E. coli.

Morphological Diversity of Dps Complex with Genomic DNA.

In response to adverse environmental factors, Escherichia coli cells actively produce Dps proteins which form ordered complexes (biocrystals) with bacterial DNA to protect the genome. The effect of biocrystallization has been described extensively in the scientific literature; furthermore, to date, the structure of the Dps-DNA complex has been established in detail in vitro using plasmid DNA. In the present work, for the first time, Dps complexes with E. coli genomic DNA were studied in vitro using cryo-electron tomography. We demonstrate that genomic DNA forms one-dimensional crystals or filament-like assemblies which transform into weakly ordered complexes with triclinic unit cells, similar to what is observed for plasmid DNA. Changing such environmental factors as pH and KCl and MgCl2 concentrations leads to the formation of cylindrical structures.

Influence of Cupric (Cu2+) Ions on the Iron Oxidation Mechanism by DNA-Binding Protein from Starved Cells (Dps) from Marinobacter nauticus.

Dps proteins (DNA-binding proteins from starved cells) are multifunctional stress defense proteins from the Ferritin family expressed in Prokarya during starvation and/or acute oxidative stress. Besides shielding bacterial DNA through binding and condensation, Dps proteins protect the cell from reactive oxygen species by oxidizing and storing ferrous ions within their cavity, using either hydrogen peroxide or molecular oxygen as the co-substrate, thus reducing the toxic effects of Fenton reactions. Interestingly, the interaction between Dps and transition metals (other than iron) is a known but relatively uncharacterized phenomenon. The impact of non-iron metals on the structure and function of Dps proteins is a current topic of research. This work focuses on the interaction between the Dps from Marinobacter nauticus (a marine facultative anaerobe bacterium capable of degrading petroleum hydrocarbons) and the cupric ion (Cu2+), one of the transition metals of greater biological relevance. Results obtained using electron paramagnetic resonance (EPR), Mössbauer and UV/Visible spectroscopies revealed that Cu2+ ions bind to specific binding sites in Dps, exerting a rate-enhancing effect on the ferroxidation reaction in the presence of molecular oxygen and directly oxidizing ferrous ions when no other co-substrate is present, in a yet uncharacterized redox reaction. This prompts additional research on the catalytic properties of Dps proteins.

Structural Insights into Iron Ions Accumulation in Dps Nanocage.

Dps (DNA-binding protein from starved cells) is well known for the structural protection of bacterial DNA by the formation of highly ordered intracellular assemblies under stress conditions. Moreover, this ferritin-like protein can perform fast oxidation of ferrous ions and subsequently accumulate clusters of ferric ions in its nanocages, thus providing the bacterium with physical and chemical protection. Here, cryo-electron microscopy was used to study the accumulation of iron ions in the nanocage of a Dps protein from Escherichia coli. We demonstrate that Fe2+ concentration in the solution and incubation time have an insignificant effect on the volume and the morphology of iron minerals formed in Dps nanocages. However, an increase in the Fe2+ level leads to an increase in the proportion of larger clusters and the clusters themselves are composed of discrete ~1-1.5 nm subunits.

The Conformation of the N-Terminal Tails of Deinococcus grandis Dps Is Modulated by the Ionic Strength.

DNA-binding proteins from starved cells (Dps) are homododecameric nanocages, with N- and C-terminal tail extensions of variable length and amino acid composition. They accumulate iron in the form of a ferrihydrite mineral core and are capable of binding to and compacting DNA, forming low- and high-order condensates. This dual activity is designed to protect DNA from oxidative stress, resulting from Fenton chemistry or radiation exposure. In most Dps proteins, the DNA-binding properties stem from the N-terminal tail extensions. We explored the structural characteristics of a Dps from Deinococcus grandis that exhibits an atypically long N-terminal tail composed of 52 residues and probed the impact of the ionic strength on protein conformation using size exclusion chromatography, dynamic light scattering, synchrotron radiation circular dichroism and small-angle X-ray scattering. A novel high-spin ferrous iron-binding site was identified in the N-terminal tails, using Mössbauer spectroscopy. Our data reveals that the N-terminal tails are structurally dynamic and alter between compact and extended conformations, depending on the ionic strength of the buffer. This prompts the search for other physiologically relevant modulators of tail conformation and hints that the DNA-binding properties of Dps proteins may be affected by external factors.

Controlled modulation of the dynamics of the Deinococcus grandis Dps N-terminal tails by divalent metals.

DNA-binding proteins from starved cells (Dps) are small multifunctional nanocages expressed by prokaryotes in acute oxidative stress conditions or during the starvation-induced stationary phase, as a bacterial defense mechanism. Dps proteins protect bacterial DNA from damage by either direct binding or by removing precursors of reactive oxygen species from solution. The DNA-binding properties of most Dps proteins studied so far are related to their unordered, flexible, N- and C-terminal extensions. In a previous work, we revealed that the N-terminal tails of Deinoccocus grandis Dps shift from an extended to a compact conformation depending on the ionic strength of the buffer and detected a novel high-spin ferrous iron center in the proximal ends of those tails. In this work, we further explore the conformational dynamics of the protein by probing the effect of divalent metals binding to the tail by comparing the metal-binding properties of the wild-type protein with a binding site-impaired D34A variant using size exclusion chromatography, dynamic light scattering, synchrotron radiation circular dichroism, and small-angle X-ray scattering. The N-terminal ferrous species was also characterized by Mössbauer spectroscopy. The results herein presented reveal that the conformation of the N-terminal tails is altered upon metal binding in a gradual, reversible, and specific manner. These observations may point towards the existence of a regulatory process for the DNA-binding properties of Dps proteins through metal binding to their N- and/or C-terminal extensions.

Bacterioferritin nanocage: Structure, biological function, catalytic mechanism, self-assembly and potential applications.

Bacterioferritin (Bfr) is a subfamily of ferritin protein family. Bfrs are composed of 24 identical subunits and self-assemble into 4-3-2-fold symmetric cage-like structure with the incorporation of 12 heme groups into twelve 2-fold symmetric binding sites between subunits. Bfr protein cage has an outer diameter of ∼12 nm and interior cavity diameter of ∼8 nm with a total of 62 pores to connect the interior cavity with the bulk solution outside the protein nanocage. In vivo, the interior cavity of Bfr can store up to ∼2700 iron atoms in the ferrihydrite-like mineral. Recent years, more and more Bfr structures have been solved, which elucidated more details about the ferroxidase center, the catalytic mechanism, the possible channels used by iron ions to access the interior cavity, the electron transfer pathway involved in the iron redox cycle, and the molecular function of the heme group. The preliminary applications of both mammalian and bacterial ferritins in drug delivery, imaging diagnosis, and nanoparticle vaccine make Bfr exploration uniquely attractive for researchers from a broad range of research fields because Bfr has advantages over ferritins in controlling the self-assembly and redesigning the subunit. In this article, we outline the structure of Bfr, review the recent progress in the molecular mechanism of Bfr to store and release iron, and focus on the self-assembly and genetic modification of Bfr nanocage. Based on the comparison between Bfr and other ferritin family members, we further discuss the potential applications of Bfr. We expect that both fundamental and applied researches on Bfr will attract broad interest in protein nanocage design, nanomedicine, precise therapy, nanoparticle vaccine, bionanotechnology, bionanoelectronics, and so on.