Role of N and C-terminal Tails in DNA Binding and Assembly in Dps: Structural Studies of Mycobacterium smegmatis Dps Deletion Mutants

Roy, Siddhartha; Saraswathi, R.; Gupta, Surbhi; Sekar, K.; Chatterji, Dipankar; Vijayan, M.

doi:10.1016/j.jmb.2007.05.004

Cited by 50 publications

(48 citation statements)

References 42 publications

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“…Thus, the LiDps molecules form an ordered array on the TEM grid, with the superstructure observed by FFT ( Figure 2A and inset), in accordance with their strong tendency to produce two-and three-dimensional (2D and 3D) crystals in vitro and in vivo [16] and consistent with the formation of free-standing 3D crystals of magnetoferritin. [12] Moreover, the iron oxide nanograins within the Dps cavity are single crystals of magnetite/maghemite (Figure 2A, inset).…”

Section: Resultssupporting

confidence: 79%

Synthesis of Iron Oxide Nanoparticles in Listeria innocua Dps (DNA‐Binding Protein from Starved Cells): A Study with the Wild‐Type Protein and a Catalytic Centre Mutant

Ceci

Chiancone

Kasyutich

et al. 2009

Chemistry A European J

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A comparative analysis of the magnetic properties of iron oxide nanoparticles grown in the cavity of the DNA-binding protein from starved cells of the bacterium Listeria innocua, LiDps, and of its triple-mutant lacking the catalytic ferroxidase centre, LiDps-tm, is presented. TEM images and static and dynamic magnetic and electron magnetic resonance (EMR) measurements reveal that, under the applied preparation conditions, namely alkaline pH, high temperature (65 degrees C), exclusion of oxygen, and the presence of hydrogen peroxide, maghemite and/or magnetite nanoparticles with an average diameter of about 3 nm are mineralised inside the cavities of both LiDps and LiDps-tm. The magnetic nanoparticles (MNPs) thus formed show similar magnetic properties, with superparamagnetic behaviour above 4.5 K and a large magnetic anisotropy. Interestingly, in the EMR spectra an absorption at half-field is observed, which can be considered as a manifestation of the quantum behaviour of the MNPs. These results indicate that Dps proteins can be advantageously used for the production of nanomagnets at the interface between molecular clusters and traditional MNPs and that the presence of the ferroxidase centre, though increasing the efficiency of nanoparticle formation, does not affect the nature and fine structure of the MNPs. Importantly, the self-organisation of MNP-containing Dps on HRTEM grids suggests that Dps-enclosed MNPs can be deposited on surfaces in an ordered fashion.

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Section: Resultssupporting

confidence: 79%

Synthesis of Iron Oxide Nanoparticles in Listeria innocua Dps (DNA‐Binding Protein from Starved Cells): A Study with the Wild‐Type Protein and a Catalytic Centre Mutant

Ceci

Chiancone

Kasyutich

et al. 2009

Chemistry A European J

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“…The few mutants that have been discovered to alter the Dps oligomerization state involved deletions of relatively large subdomains that have been implicated in other roles such as making interactions with DNA or in iron transport. 34 Therefore, the discovery of a simple double mutant that cleanly alters the oligomerization state not only provides insight into fundamentals of this system, it could also provide direction to the development of unique applications and better tools to explore the role oligomerization plays in DNA binding, 30 the design of a switchable protein container, as a control for high throughput assays for screening protein libraries for enhance oligomerization or as a first step to design a protein that can form alternative oligomerization states.…”

Section: Discussionmentioning

confidence: 99%

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Zhang

Chee

et al. 2011

Protein Science

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DNA-binding protein from starved cells (DPS), a mini-ferritin capable of self-assembling into a 12-meric nano-cage, was chosen as the basis for an alanine-shaving mutagenesis study to investigate the importance of key amino acid residues, located at symmetry-related protein-protein interfaces, in controlling protein stability and self-assembly. Nine mutants were designed through simple inspection, synthesized, and subjected to transmission electron microscopy, circular dichroism, size exclusion chromatography, and ''virtual alanine scanning'' computational analysis. The data indicate that many of these residues may be hot spot residues. Most remarkably, two residues, R83 and R133, were observed to shift the oligomerization state to~50% dimer. Based on the hypothesis that these two residues constitute a ''hot strip,'' located at the ferritin-like threefold axis, the double mutant was generated which completely shuts down detectable formation of 12-mer in solution, favoring a cooperatively folded dimer. The fact that this effect logically builds upon the single mutants emphasizes that complex self-assembly has the potential to be manipulated rationally. This study should have an impact on the fundamental understanding of the assembly of DPS protein cages specifically and protein quaternary structure in general. In addition, as there is much interest in applying these and similar systems to the templation of nano-materials and drug delivery, the ability to control this ferritin's oligomerization state and stability could prove especially valuable.

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“…In dodecameric DPS proteins, this short C-terminal helix is lost, with the C-terminal tail adopting a random coil arrangement on the outside of the particle (29), where, along with the N-terminal tail, it may play a role in modulating nonspecific interactions with DNA (54). In addition, the C-terminal tail in DPS has also been shown to play a critical role in stabilizing the dodecamer (54,55). Now, in the SsDPSL and BfDPSL dodecamers, we see that the C-terminal tail can adopt two additional conformations.…”

Section: Discussionmentioning

confidence: 99%

Characterization of the Bacteroides fragilis bfr Gene Product Identifies a Bacterial DPS-Like Protein and Suggests Evolutionary Links in the Ferritin Superfamily

Gauss¹,

Reott²,

Rocha³

et al. 2011

Journal of Bacteriology

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A factor contributing to the pathogenicity of Bacteroides fragilis, the most common anaerobic species isolated from clinical infections, is the bacterium's extreme aerotolerance, which allows survival in oxygenated tissues prior to anaerobic abscess formation. We investigated the role of the bacterioferritin-related (bfr) gene in the B. fragilis oxidative stress response. The bfr mRNA levels are increased in stationary phase or in response to O 2 or iron. In addition, bfr null mutants exhibit reduced aerotolerance, and the bfr gene product protects DNA from hydroxyl radical cleavage in vitro. Crystallographic studies revealed a protein with a dodecameric structure and greater similarity to an archaeal DNA protection in starved cells (DPS)-like protein than to the 24-subunit bacterioferritins. Similarity to the DPS-like (DPSL) protein extends to the subunit and includes a pair of conserved cysteine residues juxtaposed to a buried dimetal binding site within the four-helix bundle. Compared to archaeal DPSLs, however, this bacterial DPSL protein contains several unique features, including a significantly different conformation in the C-terminal tail that alters the number and location of pores leading to the central cavity and a conserved metal binding site on the interior surface of the dodecamer. Combined, these characteristics confirm this new class of miniferritin in the bacterial domain, delineate the similarities and differences between bacterial DPSL proteins and their archaeal homologs, allow corrected annotations for B. fragilis bfr and other dpsl genes within the bacterial domain, and suggest an evolutionary link within the ferritin superfamily that connects dodecameric DPS to the (bacterio)ferritin 24-mer. Bacteroides fragilis is a strict anaerobe that comprises approximately 1 to 2% of the normal intestinal flora in humans. While normally present as a commensal bacterium in this reducing environment, it is also the pathogenic anaerobe most frequently isolated from intra-abdominal infections, abscesses, and blood (27,57,58). Factors contributing to the pathogenesis of B. fragilis include resistance to oxidative stress and extreme aerotolerance, each of which is an important virulence factor for extraintestinal infections (61).A significant pathway for the production of reactive oxygen species (ROS) is dependent upon the presence of free Fe 2ϩ , where Fe 2ϩ and O 2 react to form the toxic superoxide anion (19, 37). Biologically, superoxide is then converted to H 2 O 2 and O 2 by the action of superoxide dismutase (69) or superoxide reductase (43). Then, via the Fenton reaction, the hydrogen peroxide may react with remaining Fe 2ϩ to produce the hydroxyl radical (HO·), the most toxic of all ROSs. Management of the intracellular iron pool is thus an important facet in the control of oxidative stress in virtually all organisms.One strategy for limiting the availability of free iron is adopted by ferritin, bacterioferritin, and DNA protection in nutrientstarved cells (DPS) protein (31,37,64). These members of...

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Role of N and C-terminal Tails in DNA Binding and Assembly in Dps: Structural Studies of Mycobacterium smegmatis Dps Deletion Mutants

Cited by 50 publications

References 42 publications

Synthesis of Iron Oxide Nanoparticles in Listeria innocua Dps (DNA‐Binding Protein from Starved Cells): A Study with the Wild‐Type Protein and a Catalytic Centre Mutant

Synthesis of Iron Oxide Nanoparticles in Listeria innocua Dps (DNA‐Binding Protein from Starved Cells): A Study with the Wild‐Type Protein and a Catalytic Centre Mutant

Rational disruption of the oligomerization of the mini‐ferritin E. coli DPS through protein‐protein interface mutation

Characterization of the Bacteroides fragilis bfr Gene Product Identifies a Bacterial DPS-Like Protein and Suggests Evolutionary Links in the Ferritin Superfamily

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