The fragile X syndrome protein represses activity-dependent translation through CYFIP1, a new 4E-BP.

Strong evidence indicates that regulated mRNA translation in neuronal dendrites underlies synaptic plasticity and brain development. The fragile X mental retardation protein (FMRP) is involved in this process; here, we show that it acts by inhibiting translation initiation. A binding partner of FMRP, CYFIP1/Sra1, directly binds the translation initiation factor eIF4E through a domain that is structurally related to those present in 4E-BP translational inhibitors. Brain cytoplasmic RNA 1 (BC1), another FMRP binding partner, increases the affinity of FMRP for the CYFIP1-eIF4E complex in the brain. Levels of proteins encoded by known FMRP target mRNAs are increased upon reduction of CYFIP1 in neurons. Translational repression is regulated in an activity-dependent manner because BDNF or DHPG stimulation of neurons causes CYFIP1 to dissociate from eIF4E at synapses, thereby resulting in protein synthesis. Thus, the translational repression activity of FMRP in the brain is mediated, at least in part, by CYFIP1.

Deciphering the Broad Antimicrobial Activity of Melaleuca alternifolia Tea Tree Oil by Combining Experimental and Computational Investigations.

Tea Tree Oil (TTO) is an essential oil obtained from the distillation of Melaleuca alternifolia leaves and branches. Due to its beneficial properties, TTO is widely used as an active ingredient in antimicrobial preparations for topical use or in cosmetic products and contains about 100 different compounds, with terpinen-4-ol, γ-terpinene and 1,8-cineole (or eucalyptol) being the molecules most responsible for its biological activities. In this work, the antimicrobial activity of whole TTO and these three major components was evaluated in vitro against fungi, bacteria and viruses. Molecular dynamics simulations were carried out on a bacterial membrane model and a Coxsackievirus B4 viral capsid, to propose an atomistic explanation of their mechanism of action. The obtained results indicate that the strong antimicrobial activity of TTO is attributable to the induction of an altered membrane functionality, mediated by the incorporation of its components within the lipid bilayer, and to a possible ability of the compounds to bind and alter the structural properties of the viral capsid.

Potential Use of Tea Tree Oil as a Disinfectant Agent against Coronaviruses: A Combined Experimental and Simulation Study.

The COVID-19 pandemic has highlighted the relevance of proper disinfection procedures and renewed interest in developing novel disinfectant materials as a preventive strategy to limit SARS-CoV-2 contamination. Given its widely known antibacterial, antifungal, and antiviral properties, Melaleuca alternifolia essential oil, also named Tea tree oil (TTO), is recognized as a potential effective and safe natural disinfectant agent. In particular, the proposed antiviral activity of TTO involves the inhibition of viral entry and fusion, interfering with the structural dynamics of the membrane and with the protein envelope components. In this study, for the first time, we demonstrated the virucidal effects of TTO against the feline coronavirus (FCoVII) and the human coronavirus OC43 (HCoV-OC43), both used as surrogate models for SARS-CoV-2. Then, to atomistically uncover the possible effects exerted by TTO compounds on the outer surface of the SARS-CoV-2 virion, we performed Gaussian accelerated Molecular Dynamics simulations of a SARS-CoV-2 envelope portion, including a complete model of the Spike glycoprotein in the absence or presence of the three main TTO compounds (terpinen-4-ol, γ-terpinene, and 1,8-cineole). The obtained results allowed us to hypothesize the mechanism of action of TTO and its possible use as an anti-coronavirus disinfectant agent.

Antifungal Effect of All-trans Retinoic Acid against Aspergillus fumigatus In Vitro and in a Pulmonary Aspergillosis In Vivo Model.

Aspergillus fumigatus is the most common opportunistic fungal pathogen and causes invasive pulmonary aspergillosis (IPA), with high mortality among immunosuppressed patients. The fungistatic activity of all-trans retinoic acid (ATRA) has been recently described in vitro We evaluated the efficacy of ATRA in vivo and its potential synergistic interaction with other antifungal drugs. A rat model of IPA and in vitro experiments were performed to assess the efficacy of ATRA against Aspergillus in association with classical antifungal drugs and in silico studies used to clarify its mechanism of action. ATRA (0.5 and 1 mM) displayed a strong fungistatic activity in Aspergillus cultures, while at lower concentrations, synergistically potentiated fungistatic efficacy of subinhibitory concentration of amphotericin B (AmB) and posaconazole (POS). ATRA also enhanced macrophagic phagocytosis of conidia. In a rat model of IPA, ATRA reduced mortality similarly to posaconazole. Fungistatic efficacy of ATRA alone and synergistically with other antifungal drugs was documented in vitro, likely by inhibiting fungal heat shock protein 90 (Hsp90) expression and Hsp90-related genes. ATRA treatment reduced mortality in a model of IPA in vivo Those findings suggest ATRA as a suitable fungistatic agent that can also reduce dosage and adverse reactions of classical antifungal drugs and add to the development of new therapeutic strategies against IPA and systemic fungal infections.

Post Zygotic, Somatic, Deletion in KERATIN 1 V1 Domain Generates Structural Alteration of the K1/K10 Dimer, Producing a Monolateral Palmar Epidermolytic Nevus.

Palmoplantar keratodermas (PPKs) are characterized by thickness of stratum corneum and epidermal hyperkeratosis localized in palms and soles. PPKs can be epidermolytic (EPPK) or non epidermolytic (NEPPK). Specific mutations of keratin 16 (K16) and keratin 1 (K1) have been associated to EPPK, and NEPPK. Cases of mosaicism in PPKs due to somatic keratin mutations have also been described in scientific literature. We evaluated a patient presenting hyperkeratosis localized monolaterally in the right palmar area, characterized by linear yellowish hyperkeratotic lesions following the Blaschko lines. No other relatives of the patient showed any dermatological disease. Light and confocal histological analysis confirmed the presence of epidermolityic hyperkeratosis. Genetic analysis performed demonstrates the heterozygous deletion NM_006121.4:r.274_472del for a total of 198 nucleotides, in KRT1 cDNA obtained by a palmar lesional skin biopsy, corresponding to the protein mutation NP_006112.3:p.Gly71_Gly137del. DNA extracted from peripheral blood lymphocytes did not display the presence of the mutation. These results suggest a somatic mutation causing an alteration in K1 N-terminal variable domain (V1). The deleted sequence involves the ISIS subdomain, containing a lysine residue already described as fundamental for epidermal transglutaminases in the crosslinking of IF cytoskeleton. Moreover, a computational analysis of the wild-type and V1-mutated K1/K10 keratin dimers, suggests an unusual interaction between these keratin filaments. The mutation taster in silico analysis also returned a high probability for a deleterious mutation. These data demonstrate once again the importance of the head domain (V1) of K1 in the formation of a functional keratinocyte cytoskeleton. Moreover, this is a further demonstration of the presence of somatic mutations arising in later stages of the embryogenesis, generating a mosaic phenotype.

In Vitro and In Silico Characterization of an Antimalarial Compound with Antitumor Activity Targeting Human DNA Topoisomerase IB.

Human DNA topoisomerase IB controls the topological state of supercoiled DNA through a complex catalytic cycle that consists of cleavage and religation reactions, allowing the progression of fundamental DNA metabolism. The catalytic steps of human DNA topoisomerase IB were analyzed in the presence of a drug, obtained by the open-access drug bank Medicines for Malaria Venture. The experiments indicate that the compound strongly and irreversibly inhibits the cleavage step of the enzyme reaction and reduces the cell viability of three different cancer cell lines. Molecular docking and molecular dynamics simulations suggest that the drug binds to the human DNA topoisomerase IB-DNA complex sitting inside the catalytic site of the enzyme, providing a molecular explanation for the cleavage-inhibition effect. For all these reasons, the aforementioned drug could be a possible lead compound for the development of an efficient anti-tumor molecule targeting human DNA topoisomerase IB.

Engineering a responsive DNA triple helix into an octahedral DNA nanostructure for a reversible opening/closing switching mechanism: a computational and experimental integrated study.

We propose an experimental and simulative approach to study the effect of integrating a DNA functional device into a large-sized DNA nanostructure. We selected, as a test bed, a well-known and characterized pH-dependent clamp-switch, based on a parallel DNA triple helix, to be integrated into a truncated octahedral scaffold. We designed, simulated and experimentally characterized two different functionalized DNA nanostructures, with and without the presence of a spacer between the scaffold and the functional elements. The experimental and simulative data agree in validating the need of a spacer for the occurrence of the pH dependent switching mechanism. The system is fully reversible and the switching can be monitored several times without any perturbation, maintaining the same properties of the isolated clamp switch in solution.

Dynamical Behavior of the Human Ferroportin Homologue from Bdellovibrio bacteriovorus: Insight into the Ligand Recognition Mechanism.

Members of the major facilitator superfamily of transporters (MFS) play an essential role in many physiological processes such as development, neurotransmission, and signaling. Aberrant functions of MFS proteins are associated with several diseases, including cancer, schizophrenia, epilepsy, amyotrophic lateral sclerosis and Alzheimer's disease. MFS transporters are also involved in multidrug resistance in bacteria and fungi. The structures of most MFS members, especially those of members with significant physiological relevance, are yet to be solved. The lack of structural and functional information impedes our detailed understanding, and thus the pharmacological targeting, of these transporters. To improve our knowledge on the mechanistic principles governing the function of MSF members, molecular dynamics (MD) simulations were performed on the inward-facing and outward-facing crystal structures of the human ferroportin homologue from the Gram-negative bacterium Bdellovibrio bacteriovorus (BdFpn). Several simulations with an excess of iron ions were also performed to explore the relationship between the protein's dynamics and the ligand recognition mechanism. The results reinforce the existence of the alternating-access mechanism already described for other MFS members. In addition, the reorganization of salt bridges, some of which are conserved in several MFS members, appears to be a key molecular event facilitating the conformational change of the transporter.

Specific miRNA and Gene Deregulation Characterize the Increased Angiogenic Remodeling of Thoracic Aneurysmatic Aortopathy in Marfan Syndrome.

Marfan syndrome (MFS) is a connective tissue disease caused by mutations in the FBN1 gene, leading to alterations in the extracellular matrix microfibril assembly and the early formation of thoracic aorta aneurysms (TAAs). Non-genetic TAAs share many clinico-pathological aspects with MFS and deregulation of some microRNAs (miRNAs) has been demonstrated to be involved in the progression of TAA. In this study, 40 patients undergoing elective ascending aorta surgery were enrolled to compare TAA histomorphological features, miRNA profile and related target genes in order to find specific alterations that may explain the earlier and more severe clinical outcomes in MFS patients. Histomorphological, ultrastructural and in vitro studies were performed in order to compare aortic wall features of MFS and non-MFS TAA. MFS displayed greater glycosaminoglycan accumulation and loss/fragmentation of elastic fibers compared to non-MFS TAA. Immunohistochemistry revealed increased CD133+ angiogenic remodeling, greater MMP-2 expression, inflammation and smooth muscle cell (SMC) turnover in MFS TAA. Cultured SMCs from MFS confirmed higher turnover and α-smooth muscle actin expression compared with non-MFS TAA. Moreover, twenty-five miRNAs, including miR-26a, miR-29, miR-143 and miR-145, were found to be downregulated and only miR-632 was upregulated in MFS TAA in vivo. Bioinformatics analysis revealed that some deregulated miRNAs in MFS TAA are implicated in cell proliferation, extracellular matrix structure/function and TGFß signaling. Finally, gene analysis showed 28 upregulated and seven downregulated genes in MFS TAA, some of them belonging to the CDH1/APC and CCNA2/TP53 signaling pathways. Specific miRNA and gene deregulation characterized the aortopathy of MFS and this was associated with increased angiogenic remodeling, likely favoring the early and more severe clinical outcomes, compared to non-MFS TAA. Our findings provide new insights concerning the pathogenetic mechanisms of MFS TAA; further investigation is needed to confirm if these newly identified specific deregulated miRNAs may represent potential therapeutic targets to counteract the rapid progression of MFS aortopathy.

Probing the Functional Topology of a pH-Dependent Triple Helix DNA Nanoswitch Family through Gaussian Accelerated MD Simulation.

The topology of a pH-dependent triple helix DNA nanoswitch family has been characterized through simulative analysis to evaluate the efficiency of the switching mechanism varying the length of the loop connecting the two strands forming the double helix portion. In detail, the system is formed by a double helix made by two six base complementary sequences, connected by one loop having an increasing number of thymidines, namely 5, 7, or 9. The triplex-forming sequence made by six bases, connected to the double helix through a constant 25 base loop, interacts at pH 5.0 through Hoogsteen hydrogen bonds with one strand of the double helical region. We demonstrate, through molecular dynamics simulation, that the thymidine loop length exerts a fine regulatory role for the stability of the triple helix structure and is critical in modulating the switching mechanism triggered by the pH increase.

Luteolin-7-O-ß-d-Glucoside Inhibits Cellular Energy Production Interacting with HEK2 in Keratinocytes.

Flavonoids have been demonstrated to affect the activity of many mammalian enzyme systems. Their functional phenolic groups are able to mediate antioxidant effects by scavenging free radicals. Molecules of this class have been found able to modulate the activity of kinases, phospholipase A2, cyclooxygenases, lipoxygenase, glutathione S-transferase, and many others. Recently, it has been demonstrated that luteolin, in the form of Luteolin-7-O-ß-d-glucoside (LUT-7G) is able to induce the keratinocyte differentiation process in vitro. This flavonoid is able to counteract the proliferative effects of IL-22/IL6 pathway by the inhibition of STAT3 activity also in vivo in a psoriatic mouse model. Observations on energy metabolism changes of differentiating cells led us to perform a complete metabolomics analysis using human primary keratinocytes treated with LUT-7G. Our results show that LUT-7G, is not only able to impair the nuclear translocation of STAT3, but it also blocks the energy metabolism pathway, depressing the glycolytic and Krebs pathway by the inhibition of hexokinase 2 activity. These data confirm that LUT-7G can be proposed as a potential candidate for the treatment of inflammatory and proliferative diseases, but its role as a hexokinase 2 (HEK2) inhibitor opens new perspectives in nutritional science, and especially in cancer therapy, in which the inhibition of the Warburg effect could be relevant.

Succinic semialdehyde dehydrogenase deficiency: The combination of a novel ALDH5A1 gene mutation and a missense SNP strongly affects SSADH enzyme activity and stability.

Succinic semialdehyde dehydrogenase deficiency (SSADHD) is a rare autosomal recessive metabolic disorder of GABA catabolism. SSADH is a mitochondrial homotetrameric enzyme encoded by ALDH5A1 gene. We report the molecular characterization of ALDH5A1 gene in an Italian SSADHD patient, showing heterozygosity for four missense mutations: c.526G>A (p.G176R), c.538C>T (p.H180Y), c.709G>T (p.A237S) and c.1267A>T (p.T423S), the latter never described so far. The patient inherited c.526A in cis with c.538T from the mother and c.709T in cis with c.1267T from the father. To explore the effects of the two allelic arrangements on SSADH activity and protein level, wild type, single or double mutated cDNA constructs were expressed in a cell system. The p.G176R change, alone or in combination with p.H180Y, causes the abolishment of enzyme activity. Western blot analysis showed a strongly reduced amount of the p.176R-p.180Y double mutant protein, suggesting increased degradation. Indeed, in silico analyses confirmed high instability of this mutant homotetramer. Enzyme activity relative to the other p.423S-p.237S double mutant is around 30% of wt. Further in silico analyses on all the possible combinations of mutant monomers suggest the lowest stability for the tetramer constituted by p.176R-p.180Y monomers and the highest stability for that constituted by p.237S-p.423S monomers. The present study shows that when a common SNP, associated with a slight reduction of SSADH activity, is inherited in cis with a mutation showing no consequences on the enzyme function, the activity is strongly affected. In conclusion, the peculiar arrangement of four missense mutations occurring in this patient is responsible for the SSADHD phenotype.

Molecular dynamics characterization of the SAMHD1 Aicardi-Goutières Arg145Gln mutant: structural determinants for the impaired tetramerization.

Aicardi-Goutières syndrome, a rare genetic disorder characterized by calcification of basal ganglia, results in psychomotor delays and epilepsy states from the early months of children life. This disease is caused by mutations in seven different genes encoding proteins implicated in the metabolism of nucleic acids, including SAMHD1. Twenty SAMHD1 gene variants have been discovered and in this work, a structural characterization of the SAMHD1 Aicardi-Goutières Arg145Gln mutant is reported by classical molecular dynamics simulation. Four simulations have been carried out and compared. Two concerning the wild-type SAMHD1 form in presence and absence of cofactors, in order to explain the role of cofactors in the SAMHD1 assembly/disassembly process and, two concerning the Arg145Gln mutant, also in presence and absence of cofactors, in order to have an accurate comparison with the corresponding native forms. Results show the importance of native residue Arg145 in maintaining the tetramer, interacting with GTP cofactor inside allosteric sites. Replacement of arginine in glutamine gives rise to a loosening of GTP-protein interactions, when cofactors are present in allosteric sites, whilst in absence of cofactors, the occurrence of intra and inter-chain interactions is observed in the mutant, not seen in the native enzyme, making energetically unfavourable the tetramerization process.

Salmonella enterica serovar Typhimurium growth is inhibited by the concomitant binding of Zn(II) and a pyrrolyl-hydroxamate to ZnuA, the soluble component of the ZnuABC transporter.

BACKGROUND: Under conditions of Zn(II) deficiency, the most relevant high affinity Zn(II) transport system synthesized by many Gram-negative bacteria is the ZnuABC transporter. ZnuABC is absent in eukaryotes and plays an important role in bacterial virulence. Consequently, ZnuA, the periplasmic component of the transporter, appeared as a good target candidate to find new compounds able to contrast bacterial growth by interfering with Zn(II) uptake. METHODS: Antibacterial activity assays on selected compounds from and in-house library against Salmonella enterica serovar Typhimurium ATCC14028 were performed. The X-ray structure of the complex formed by SeZnuA with an active compound was solved at 2.15Å resolution. RESULTS: Two di-aryl pyrrole hydroxamic acids differing in the position of a chloride ion, RDS50 ([1-[(4-chlorophenyl)methyl]-4-phenyl-1H-pyrrol-3-hydroxamic acid]) and RDS51 (1-[(2-chlorophenyl)methyl]-4-phenyl-1H-pyrrol-3-hydroxamic acid) were able to inhibit Salmonella growth and its invasion ability of Caco-2 cells. The X-ray structure of SeZnuA containing RDS51 revealed its presence at the metal binding site concomitantly with Zn(II) which is coordinated by protein residues and the hydroxamate moiety of the compound. CONCLUSIONS: Two molecules interfering with ZnuA-mediated Zn(II) transport in Salmonella have been identified for the first time. The resolution of the SeZnuA-RDS51 X-ray structure revealed that RDS51 is tightly bound both to the protein and to Zn(II) thereby inhibiting its release. These features pave the way to the rational design of new Zn(II)-binding drugs against Salmonella. GENERAL SIGNIFICANCE: The data reported show that targeting the bacterial ZnuABC transporter can represent a good strategy to find new antibiotics against Gram-negative bacteria.

Multiple molecular dynamics simulations of human LOX-1 and Trp150Ala mutant reveal the structural determinants causing the full deactivation of the receptor.

Multiple classical molecular dynamics simulations have been applied to the human LOX-1 receptor to clarify the role of the Trp150Ala mutation in the loss of binding activity. Results indicate that the substitution of this crucial residue, located at the dimer interface, markedly disrupts the wild-type receptor dynamics. The mutation causes an irreversible rearrangement of the subunits interaction pattern that in the wild-type protein allows the maintaining of a specific symmetrical motion of the monomers. The subunits dislocation determines a loss of linearity of the arginines residues composing the basic spine and a consequent alteration of the long-range electrostatic attraction of the substrate. Moreover, the anomalous subunits arrangement observed in the mutated receptor also affects the integrity of the hydrophobic tunnel, actively involved in the short-range hydrophobic recognition of the substrate. The combined effect of these structural rearrangements generates the impairing of the receptor function.

Simulative and Experimental Characterization of a pH-Dependent Clamp-like DNA Triple-Helix Nanoswitch.

Here we couple experimental and simulative techniques to characterize the structural/dynamical behavior of a pH-triggered switching mechanism based on the formation of a parallel DNA triple helix. Fluorescent data demonstrate the ability of this structure to reversibly switch between two states upon pH changes. Two accelerated, half microsecond, MD simulations of the system having protonated or unprotonated cytosines, mimicking the pH 5.0 and 8.0 conditions, highlight the importance of the Hoogsteen interactions in stabilizing the system, finely depicting the time-dependent disruption of the hydrogen bond network. Urea-unfolding experiments and MM/GBSA calculations converge in indicating a stabilization energy at pH 5.0, 2-fold higher than that observed at pH 8.0. These results validate the pH-controlled behavior of the designed structure and suggest that simulative approaches can be successfully coupled with experimental data to characterize responsive DNA-based nanodevices.

A molecular dynamics simulation study decodes the early stage of the disassembly process abolishing the human SAMHD1 function.

The human sterile alpha motif SAM and HD domain-containing protein 1 (SAMHD1) restricts in non-cycling cells type the infection of a large range of retroviruses including HIV-1, reducing the intracellular pool concentration of deoxynucleoside triphosphates (dNTPs) required for the reverse transcription of the viral genome. The enzyme is in equilibrium between different forms depending on bound cofactors and substrate. In this work, two SAMHD1 three-dimensional models have been investigated through classical molecular dynamics simulation, to define the role of cofactors and metal ions in the association of the tetrameric active form. A detailed analysis of the inter-subunit interactions, taking place at the level of helix 13, indicates that removal of metal ions and cofactors induces an asymmetric loosening of the monomer-monomer interface leading to the formation of a loose tetramer where the two dimeric interfaces are weakened in different way.

SSADH deficiency in an Italian family: a novel ALDH5A1 gene mutation affecting the succinic semialdehyde substrate binding site.

SSADH deficiency (SSADHD) is a rare autosomal recessively inherited metabolic disorder. It is associated with mutations of ALDH5A1 gene, coding for the homotetrameric enzyme SSADH. This enzyme is involved in γ-aminobutyric acid (GABA) catabolism, since it oxidizes succinic semialdehyde (SSA) to succinate. Mutations in ALDH5A1 gene result in the abnormal accumulation of γ-hydroxybutyrate (GHB), which is pathognomonic of SSADHD. In the present report, diagnosis of SSADHD in a three-month-old female was achieved by detection of high levels of GHB in urine. Sequence analysis of ALDH5A1 gene showed that the patient was a compound heterozygote for c.1226G > A (p.G409D) and the novel missense mutation, c.1498G > C (p.V500 L). By ALDH5A1 gene expression in transiently transfected HEK293 cells and enzyme activity assays, we demonstrate that the p.V500 L mutation, despite being conservative, produces complete loss of enzyme activity. In silico protein modelling analysis and evaluation of tetramer destabilizing energies suggest that structural impairment and partial occlusion of the access channel to the active site affect enzyme activity. These findings add further knowledge on the missense mutations associated with SSADHD and the molecular mechanisms underlying the loss of the enzyme activity.

A new nitrobenzoxadiazole-based GSTP1-1 inhibitor with a previously unheard of mechanism of action and high stability.

CONTEXT: The nitrobezoxadiazole derivative NBDHEX is a potent inhibitor of glutathione transferase P1-1 (GSTP1-1) endowed with outstanding anticancer activity in different tumor models. OBJECTIVE: To characterize by in vitro biochemical and in silico studies the NBDHEX analogues named MC2752 and MC2753. MATERIALS AND METHODS: Synthesis of MC2752 and MC2753, biochemical assays and in silico docking and normal-mode analyses. RESULTS: The presence of a hydrophobic moiety in the side chain of MC2753 confers unique features to this molecule. Unlike its parent drug NBDHEX, MC2753 does not require GSH to trigger the dissociation of the complex between GSTP1-1 and TRAF2, and displays high stability towards the nucleophilic attack of the tripeptide under physiological conditions. DISCUSSION AND CONCLUSION: MC2753 may represent a lead compound for the development of novel GSTP1-1 inhibitors not affected in their anticancer action by fluctuations of cellular GSH levels, and characterized by an increased half-life in vivo.

Intrinsic Dynamics Analysis of a DNA Octahedron by Elastic Network Model.

DNA is a fundamental component of living systems where it plays a crucial role at both functional and structural level. The programmable properties of DNA make it an interesting building block for the construction of nanostructures. However, molecular mechanisms for the arrangement of these well-defined DNA assemblies are not fully understood. In this paper, the intrinsic dynamics of a DNA octahedron has been investigated by using two types of Elastic Network Models (ENMs). The application of ENMs to DNA nanocages include the analysis of the intrinsic flexibilities of DNA double-helices and hinge sites through the calculation of the square fluctuations, as well as the intrinsic collective dynamics in terms of cross-collective map calculation coupled with global motions analysis. The dynamics profiles derived from ENMs have then been evaluated and compared with previous classical molecular dynamics simulation trajectories. The results presented here revealed that ENMs can provide useful insights into the intrinsic dynamics of large DNA nanocages and represent a useful tool in the field of structural DNA nanotechnology.