Manganese diboride (MnB2) possesses an unusually high enthalpy of combustion, especially compared to other transition metal diborides (TMB). Previous work attempted to shed light on the mechanism underlying the energetics of MnB2 through the lens of overcoordination. Deeper investigation of the electronic structure and properties of MnB2 revealed potential spin crossover (SCO) behavior. The dynamic coexistence of the stable high spin (HS) state and the unstable low spin (LS) state potentially explains the material’s previously observed metastability. Applying this theory, the energetic output was tempered through the combustion of MnB2 in the presence of a magnetic field to spin trap the electrons into the more stable HS state. This opens the door to modulation of energetic output and improved storage safety of solid state energetic materials.

The ideal aeronautical solid-state fuel should possess a high gravimetric heat of combustion (more energy for less weight) and a high volumetric heat of combustion (more room for mission-critical items). In this work, manganese diboride (MnB2) demonstrates a high gravimetric heat of combustion of 39.26 kJ/g and the highest volumetric heat of combustion of any known fuel of 208.08 kJ/cm3. When compared to the currently used fuel in Space Shuttle rocket boosters and the Space Launch System, aluminum metal, MnB2 represents a 26% increase in gravimetric heat of combustion and a 148% increase in volumetric heat of combustion. Surprisingly, the local topology of the inner coordination sphere controls energetic output. A model cluster system analyzed by density functional theory shows that the local environment can contribute to the bulk properties even without physical manifestations in the periodic structure. This high enthalpic performance comes from the metastability of MnB2 and demonstrates that transition metals, typically shunned as solid-state fuels, can store potential energy from their high-temperature synthesis through 'overcoordination' and violation of their valence shell.

Positron decays of proton-rich nuclides exhibit large Q values, producing complex cascades which frequently involve various radiations, including protons and γ rays. Often, only one of the two is measured in a single experiment, limiting the accuracy and completeness of the decay scheme. An example is ^31Cl, for which protons and γ rays have been measured separately in detail but never with substantial sensitivity to proton-γ coincidences. We provide detailed measurements of ^31Cl β-delayed proton decay including β-p-γ sequences, extract spectroscopic information on ^31S excited states as well as their β^+ feedings, and compare to shell-model calculations. A fast fragmented beam of ^31Cl provided by the National Superconducting Cyclotron Laboratory (NSCL) was deposited in the Gaseous Detector with Germanium Tagging (GADGET) system. GADGET's gas-filled Proton Detector was used to detect β-delayed protons, and the Segmented Germanium Array (SeGA) was used to detect β-delayed γ rays. As many as 20 previously unobserved β-delayed proton transitions are reported, most of which populate excited states of ^30P. The first detailed ^31Cl(βp γ)^30P decay scheme is presented, including updated β-delayed proton energies and intensities, as well as several new ^31S levels. Improved agreement is found with theoretical calculations of the Gamow-Teller strengths B(GT) for ^31S excitation energies 7.5 < E_x < 9.5 MeV. The present work demonstrates that the ability to detect β-delayed protons and γ rays in coincidence is essential for accurate positron decay schemes to compare with nuclear structure theory. This phenomenon for β-delayed protons resembles the pandemonium effect originally introduced for β-delayed γ rays.

Nucleic acid structures are stabilized by both base pairing and base stacking. While energetics of base pairing interactions are relatively well established, our understanding of the energetic contributions of base stacking remain incomplete. Here, we use a combination of single-molecule and computational biophysics approaches to investigate the effect of strand polarity on base-stacking energetics. We designed pairs of DNA constructs with reversed stacking polarities at nick sites, along with corresponding no-stack controls to isolate stacking contributions. Performing single-molecule force-clamp assays with a Centrifuge Force Microscope (CFM), we observed polarity-dependent differences in stacking energetics. These differences were most pronounced in purine-purine and certain purine-pyrimidine interactions. Notably, a 5 ' purine stacked on a 3 ' pyrimidine was generally more stable than the reverse polarity. We employed molecular dynamics (MD) simulations to observe stacking interfaces in the DNA constructs. The simulations were qualitatively consistent with our experiments, and showed positional differences between opposite polarity stacking pairs, giving some insight into the origin of these polarity differences. Overall, these results demonstrate that base polarity can modulate stacking stability and should be considered when designing short duplex regions such as overhangs in molecular biology and biotechnology applications.

DNA nanotechnology uses the programmable assembly of DNA to create nanoscale objects. Recent work from our laboratory suggested that terminal stacking interactions between adjacent strands could be a design parameter for DNA nanotechnology. Here, we explore that idea by creating DNA tetrahedra with sticky ends containing identical base pairing interactions but different stacking interactions. Testing all 16 stacking combinations, we found that the melting temperature of DNA tetrahedra varied by up to 10 °C from altering a single base stack in the design. We also show that a 4 bp sticky end with weak stacking does not form stable tetrahedra, while strengthening the stacks confers high stability with a 46.8 ± 1.2 °C melting temperature, comparable to that of a 6 bp sticky end with weak stacking (49.7 ± 2.9 °C). The results likely apply to other DNA nanostructures and suggest that stacking interactions play a role in the formation and stability of DNA nanostructures.

The influence of aqueous solutions of 2-(tetrafluoro(trifluoromethyl)-λ6-sulfanyl-ethan-1-ol (CF3SF4-ethanol) and 2,2,2-trifluoroethanol (TFE) on the secondary structure of melittin was studied using circular dichroism (CD) and molecular dynamics (MD) simulations. In water, melittin transitions into a random coil. However, upon addition of even as little as 1% by volume of CF3SF4-ethanol, the secondary structure of melittin stabilizes as a helix. Contrarily, the addition of 40% by volume of TFE is required for the greatest helicity. Fluoroalcohols stabilize melittin's hydrophobic side chain residues, thereby enhancing the helical structure. Locally alcohol concentrations approach nearly 70-90% in the near vicinity of the hydrophobic side chains increasing hydrophobic interactions and reducing water-peptide hydrogen bonding. Using the molecular mechanics-Poisson Boltzmann surface area method (MMPBSA), the free energy of binding between the peptide and fluoroalcohols highlighted the role of nonpolar residues in stabilizing the secondary structure. Secondary structure content analysis (SESCA) validated the simulation results, confirming CF3SF4-ethanol as an effective, eco-friendly enhancer of helicity at low concentrations. The far UV circular dichroism (CD) spectrum of melittin in solutions containing TFE corroborates previous findings and likewise affirms that the addition of CF3SF4-ethanol to an aqueous solution can enhance helicity. The agreement between the experimental and calculated helicities highlights the potential of CF3SF4-ethanol. This study offers insights into peptide stabilization by fluoroalcohols, with implications for peptide-based therapeutic design.

Post-transcriptional modifications in RNA can significantly impact their structure and function. In particular, transfer RNAs (tRNAs) are heavily modified, with around 100 different naturally occurring nucleotide modifications contributing to codon bias and decoding efficiency. Here, we describe our efforts to investigate the impact of RNA modifications on the structure and stability of tRNA Phenylalanine (tRNA Phe ) from S. cerevisiae using molecular dynamics (MD) simulations. Through temperature replica exchange MD (T-REMD) studies, we explored the unfolding pathway to understand how RNA modifications influence the conformational dynamics of tRNA Phe , both in the presence and absence of magnesium ions (Mg 2+ ). We observe that modified nucleotides in key regions of the tRNA establish a complex network of hydrogen bonds and stacking interactions which is essential for tertiary structure stability of the tRNA. Furthermore, our simulations show that modifications facilitate the formation of ion binding sites on the tRNA. However, high concentrations of Mg 2+ ions can stabilize the tRNA tertiary structure in the absence of modifications. Our findings illuminate the intricate interactions between modifications, magnesium ions, and RNA structural stability.

ABSTRACT microRNAs (miRNAs) regulate target mRNA expression post-transcriptionally through their association with Argonaute 2 (AGO2) proteins. Predicting the efficiency of mRNA repression by miRNA has been limited by our comprehension of the structure-function relationship within miRNA binding sites. Using a combination of EMSA, luciferase reporter assays, and structural probing, we investigated the interaction between the human tumour suppressor miR-34a and 12 mRNA targets. Comparison of direct RNA:RNA interactions and those within the functional AGO2 protein revealed that the isolated mRNA:miRNA duplex serves as a strong predictor for duplex affinity and structure within AGO2. Our findings reveal that AGO2 has a bidirectional capacity to modulate affinity; weakening tight RNA:RNA binders while strengthening weak ones. We identified three distinct structural groups that form upon miR-34a binding and reveal a novel structural group that exhibits a guide strand bulge. MD simulations indicate a conceivable fit of this miRNA-bulge structure within AGO2. Our results demonstrate that the structural characteristics of mRNA:miRNA duplexes could serve as contributing determinants of repression efficacy.

Alternative splicing (AS) of Exon 11 of the Insulin Receptor ( INSR ) is highly regulated and disrupted in several human disorders. To better understand INSR exon 11 AS regulation, splicing activity of an INSR exon 11 minigene reporter was measured across a gradient of the AS regulator muscleblind-like 1 protein (MBNL1). The RNA-binding protein Fox-1 (RBFOX1) was added to determine its impact on MBNL1-regulated splicing. The role of the RBFOX1 UGCAUG binding site within intron 11 was assessed across the MBNL1 gradient. Mutating the UGCAUG motif inhibited RBFOX1 regulation of exon 11 and had the unexpected effect of reducing MBNL1 regulation of this exon. Molecular dynamics simulations showed that exon 11 and the adjacent RNA adopts a dynamically stable conformation. Mutation of the RBFOX1 binding site altered RNA structure and dynamics, while a mutation that created an optimal MBNL1 binding site at the RBFOX1 site shifted the RNA back to wild type. An antisense oligonucleotide (ASO) was used to confirm the structure in this region of the pre-mRNA. This example of intronic mutations shifting pre-mRNA structure and dynamics to modulate splicing suggests RNA structure and dynamics should be taken into consideration for AS regulation and therapeutic interventions targeting pre-mRNA.

A central question in biology is how RNA sequence changes influence dynamic conformational changes during cotranscriptional folding. Here we investigated this question through the study of transcriptional fluoride riboswitches, non-coding RNAs that sense the fluoride anion through the coordinated folding and rearrangement of a pseudoknotted aptamer domain and a downstream intrinsic terminator expression platform. Using a combination of Escherichia coli RNA polymerase in vitro transcription and cellular gene expression assays, we characterized the function of mesophilic and thermophilic fluoride riboswitch variants. We showed that only variants containing the mesophilic pseudoknot function at 37°C. We next systematically varied the pseudoknot sequence and found that a single wobble base pair is critical for function. Characterizing thermophilic variants at 65°C through Thermus aquaticus RNA polymerase in vitro transcription showed the importance of this wobble pair for function even at elevated temperatures. Finally, we performed all-atom molecular dynamics simulations which supported the experimental findings, visualized the RNA structure switching process, and provided insight into the important role of magnesium ions. Together these studies provide deeper insights into the role of riboswitch sequence in influencing folding and function that will be important for understanding of RNA-based gene regulation and for synthetic biology applications.

ABSTRACT Understanding the structural dynamics of how small molecule ligand recognize its RNA binding pocket is always a crucial determinant in pharmaceutical research. Molecular dynamics (MD) simulation is often used to interpretate this process at atomic resolution. However, the insurmountable high energy barriers in the binding pathway results in the nonergodic dynamics for unbiased MD sampling. To address this limitation, we applied well-tempered metadynamics coupled with upper wall restrain in this work, therefore providing an novel modeling approach for sampling the multiple state transitions during this binding process and probing the most energy favorable binding modes through two-dimensional free energy landscape reconstructed by incorporating couple possible hydrogen binding interactions between small molecule ligand and its RNA binding pocket as collective variables (CVs). Our computational predictions of binding modes for all five cases studied are in quantitative agreement with structures solved by X-ray crystallography or NMR with RMSD less than 2.0 Å. In addition, we presented the first molecular dynamics binding pathway and binding mechanism for the three cases of in vitro selected RNA aptamer. Our study demonstrated that metadynamics can be applied to effectively sampling state transitions of ligand binding events. By coupling with upper wall restrain, we have enabled fast free energy profile calculation and binding mode prediction for small molecule-RNA binding process, facilitating RNA-ligand binding investigation. This method therefore could be much-needed in computer-aided drug design pipelines of RNA-targeted small molecule compounds. Abstract Figure

A new potential termed the Fluctuating Density (FD) model is presented as an alternative to traditional fixed partial-charge force fields for describing RNA nucleobase interactions within a molecular mechanics framework. Instead of atom-centered point charges, we take inspiration from fluctuating charge models and use a site-centered density representation for electron distributions to account for charge penetration effects present in both frozen and polarizable electrostatic interactions. A parameterization procedure is established to fit the FD model against energy decomposition analysis (EDA) for density functional theory (DFT) computations of both hydrogen-bonded and stacked RNA base pairs. Additionally, the FD model's ability to replicate intermolecular interactions of NMR resolved base pairs from the RSCB Protein Data Bank, as well as a comparison against the CHARMM Drude oscillator and AMOEBA force fields, is also presented. We find that charge penetration effects, when used as a cornerstone for parameterization, have strong influences on the remaining force field energy terms such as polarization and charge transfer. Furthermore, the FD model is able to produce polarization energetic contributions from electron density changes of similar magnitude as predicted by DFT computations.

The ability to create stimuli-responsive DNA nanostructures has played a prominent role in dynamic DNA nanotechnology. Primary among these is the process of toehold-based strand displacement, where a nucleic acid molecule can act as a trigger to cause conformational changes in custom-designed DNA nanostructures. Here, we add another layer of control to strand displacement reactions through a 'toehold clipping' process. By designing DNA complexes with a photocleavable linker-containing toehold or an RNA toehold, we show that we can use light (UV) or enzyme (ribonuclease) to eliminate the toehold, thus preventing strand displacement reactions. We use molecular dynamics simulations to analyze the structural effects of incorporating a photocleavable linker in DNA complexes. Beyond simple DNA duplexes, we also demonstrate the toehold clipping process in a model DNA nanostructure, by designing a toehold containing double-bundle DNA tetrahedron that disassembles when an invading strand is added, but stays intact after the toehold clipping process even in the presence of the invading strand. This work is an example of combining multiple physical or molecular stimuli to provide additional remote control over DNA nanostructure reconfiguration, advances that hold potential use in biosensing, drug delivery or molecular computation.

Molecular dynamics (MD) simulations have become increasingly powerful and can now describe the folding/unfolding of small biomolecules in atomic detail. However, a major challenge in MD simulations is to represent the complex energy landscape of biomolecules using a small number of reaction coordinates. In this study, we investigate the folding pathways of an RNA tetraloop, gcGCAAgc, using five classical MD simulations with a combined simulation time of approximately 120 μs. Our approach involves analyzing the tetraloop dynamics, including the folding transition state ensembles, using the energy landscape visualization method (ELViM). The ELViM is an approach that uses internal distances to compare any two conformations, allowing for a detailed description of the folding process without requiring root mean square alignment of structures. This method has previously been applied to describe the energy landscape of disordered β-amyloid peptides and other proteins. The ELViM results in a non-linear projection of the multidimensional space, providing a comprehensive representation of the tetraloop's energy landscape. Our results reveal four distinct transition-state regions and establish the paths that lead to the folded tetraloop structure. This detailed analysis of the tetraloop's folding process has important implications for understanding RNA folding, and the ELViM approach can be used to study other biomolecules.

Base stacking interactions between adjacent bases in DNA and RNA are important for many biological processes and in biotechnology applications. Previous work has estimated stacking energies between pairs of bases, but contributions of individual bases has remained unknown. Here, we use a Centrifuge Force Microscope for high-throughput single molecule experiments to measure stacking energies between adjacent bases. We found stacking energies strongest between purines (G|A at -2.3 +/- 0.2 kcal/mol) and weakest between pyrimidines (C|T at -0.5 +/- 0.1 kcal/mol). Hybrid stacking with phosphorylated, methylated, and RNA nucleotides had no measurable effect, but a fluorophore modification reduced stacking energy. We experimentally show that base stacking can influence stability of a DNA nanostructure, modulate kinetics of enzymatic ligation, and assess accuracy of force fields in molecular dynamics simulations. Our results provide insights into fundamental DNA interactions that are critical in biology and can inform design in biotechnology applications. In this work, the authors use a centrifuge force microscope for high-throughput single-molecule experiments to elucidate stacking energies between individual bases of DNA.

2-Tetrafluoro(trifluoromethyl)-λ6-sulfanylethan-1-ol (CF3SF4-ethanol) combines the polar hydrophobicity of tetrafluoro(trifluoromethyl)-λ6-sulfanyl (CF3SF4) group with the polarity of simple alcohols. The properties of aqueous solutions of the well-known fluorinated alcohols 2,2,2-trifluoroethanol (TFE) and 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) were compared with those of aqueous solutions of the novel CF3SF4-ethanol. Those properties were computed using all atom molecular dynamics simulations with OPLS-compatible parameters. DFT ab initio calculations were used to accurately describe the nonsymmetrical, hypervalent sulfur in CF3SF4-ethanol. Although the molecular and conformational characteristics of CF3SF4-ethanol are like those of both TFE and HFIP, the greater hydrophobicity and lower polarity of CF3SF4-ethanol resulted in solution phase aggregation at a much lower concentration. The properties computed for TFE and HFIP in this work were consistent with published computational and experimental studies. CF3SF4-ethanol is predicted to be environmentally benign and hence an excellent green solvent candidate while possessing many of the same properties as TFE or HFIP.

RNA is critical to a broad spectrum of biological and viral processes. This functional diversity is a result of their dynamic nature; the variety of three-dimensional structures that they can fold into; and a host of post-transcriptional chemical modifications. While there are many experimental techniques to study the structural dynamics of biomolecules, molecular dynamics simulations (MDS) play a significant role in complementing experimental data and providing mechanistic insights. The accuracy of the results obtained from MDS is determined by the underlying physical models i.e., the force-fields, that steer the simulations. Though RNA force-fields have received a lot of attention in the last decade, they still lag compared to their protein counterparts. The chemical diversity imparted by the RNA modifications adds another layer of complexity to an already challenging problem. Insight into the effect of RNA modifications upon RNA folding and dynamics is lacking due to the insufficiency or absence of relevant experimental data. This review provides an overview of the state of MDS of modified RNA, focusing on the challenges in parameterization of RNA modifications as well as insights into relevant reference experiments necessary for their calibration.

To extend the approach of using RNA aptamers as transient protective groups for the synthesis of novel small-molecule drug derivatives from the existing aminoglycosides, we incorporated 2'-5' phosphodiester backbone modification in a structurally known neomycin RNA aptamer and studied the binding of a series of aminoglycosides using isothermal calorimetry (ITC) and molecular dynamics (MD) simulation. Experimental characterization of amikacin, a commercially available and widely used aminoglycoside for treating bacterial infections, shows that the aptamer A1 with a 2'-5' linkage between G15 and U16 exhibits a sevenfold increase in binding affinity with a lower binding energy compared to the native aptamer. Molecular dynamics (MD) simulation studies rationalize that this noncanonical linkage generates a narrower binding pocket by creating a superspiral RNA helical structure, which improves the ligand's fit in the binding pocket. These results provide new insights into applying 2'-5' linkages to diversify functional RNA aptamers as noncovalent protective groups in the synthesis of aminoglycoside derivatives, which can be further extended to other current drug molecules and complex natural compounds to make new pools of drug candidates more efficiently.

G-quadruplexes (GQs) are of particular biological significance due to their widespread role in various biological processes such as transcription, translation, and replication. These guanine rich structures are energetically stable and fold into helices with several stacked guanine tetrads that can form up to four strands. Due to their abundance in the genome and their unique structural character, GQs have emerged as a new class of molecular targets for drug development and important biotechnological applications such as in the development of biosensors and biomaterials.

The discovery of layered materials with potentially unique electrical and chemical properties has become a major focus of materials research in the past decade. II-VI layered hybrids (LHs) are a family of ligand-protected layered materials capable of isolation in few-layer form and possess emissive and electronic properties of potential relevance to semiconductor device technologies. We showed previously that, akin to black phosphorus (BP) and transition metal dichalcogenides (TMDCs), II-VI LHs are sensitive to ambient atmospheric conditions. However, the causes for degradation of these ligand-protected materials remain unclear. Using ZnSe-based LHs, we show herein that the stability of these materials is related to the length and chemistry of the organic ligands coordinated to the LH surfaces. Furthermore, exposure to isotopically enriched H218O and 18O2 reveals that H2O and O2 are both reactants contributing to ZnSe-LH degradation. An H2O-initiated degradation pathway is proposed and is supported by density functional theory (DFT) calculations. Our findings contribute to the discovery of protection strategies for layered materials and elucidate a degradation pathway that may also be applicable to other layered materials.