Nicotinamide mononucleotide supplementation mitigates osteopenia induced by modeled microgravity in rats.

Exposure to weightlessness causes severe osteopenia, resulting in raised fracture risk. The current study aimed to investigate whether nicotinamide mononucleotide (NMN) supplementation protected against the osteopenia in hindlimb unloading (HLU) rats in vivo and modeled microgravity-induced osteoblastic dysfunction in vitro. The 3-mo-old rats were exposed to HLU and intragastrically administered NMN every 3 days (500 mg/kg body weight) for 4 weeks. NMN supplementation mitigated HLU-induced bone loss, evidenced by greater bone mass and biomechanical properties and better trabecular bone structure. NMN supplementation mitigated HLU-induced oxidative stress, evidenced by greater levels of nicotinamide adenine dinucleotide and activities of superoxide dismutase 2 and lesser malondialdehyde levels. Modeled microgravity stimulation using rotary wall vessel bioreactor in MC3T3-E1 cells inhibited osteoblast differentiation, which was reversed by NMN treatment. Furthermore, NMN treatment mitigated microgravity-induced mitochondrial impairments, evidenced by lesser reactive oxygen species generation and greater adenosine triphosphate production, mtDNA copy number, and activities of superoxide dismutase 2 and Complex I and II. Additionally, NMN promoted activation of AMP-activated protein kinase (AMPK), evidenced by greater AMPKα phosphorylation. Our research suggested that NMN supplementation attenuated osteoblastic mitochondrial impairment and mitigated osteopenia induced by modeled microgravity.

Modeling the syn-cycle in the light activated opening of the channelrhodopsin-2 ion channel.

The ion channel of channelrhodopsin-2 (ChR2) is activated by absorbing light. The light stimulates retinal to isomerize to start the photocycle. There are two pathways for photocycles, which are caused by isomerization of the retinal from all-trans, 15-anti to 13-cis, 15-anti in the dark-adapted state (anti-cycle) and from 13-cis, 15-syn to all-trans, 15-syn in the light-adapted state (syn-cycle). In this work, the structure of the syn-cycle intermediate and mechanism of channel opening were studied by molecular dynamics (MD) and steered molecular dynamics (SMD) simulations. Due to the lack of crystal structure of intermediates in the syn-cycle of ChR2, the intermediate models were constructed from the homologous intermediates in the anti-cycle. The isomerization of retinal was shown to cause the central gate (CG) hydrogen bond network to rearrange, cutting the link between TM2 and TM7. TM2 is moved by the intrahelical hydrogen bond of E90 and K93, and induced the intracellular gate (ICG) to expand. The ion penetration pathway between TM1, TM2, TM3 and TM7 in the P500* state was observed by MD simulations. However, this channel is not fully opened compared with the homologous P500 state in the anti-cycle. In addition, the protons on Schiff bases were found to be unable to form hydrogen bonds with the counter residues (E123 and D253) in the P500* state, preventing an evolution of the P500* state to a P390-like state in the syn-cycle.

The effect on ion channel of different protonation states of E90 in channelrhodopsin-2: a molecular dynamics simulation.

Channelrhodopsin-2 (ChR2) is a cationic channel protein that has been extensively studied in optogenetics. The ion channel is opened via a series of proton transfers and H-bond changes during the photocycle but the detailed mechanism is still unknown. Molecular dynamics (MD) simulations with enhanced sampling were performed on the dark-adapted state (i.e., D470) and two photocycle intermediates (P1 500 and P2 390) to study the proton transfer path of the Schiff base and the subsequent conformational changes. The results suggest there are two possible proton transfer pathways from the Schiff base to proton acceptors (i.e., E123 or D253), depending on the protonation of E90. If E90 is protonated in the P1 500 state, the proton on the Schiff base will transfer to E123. The polyene chain of 13-cis retinal tilts and opens the channel that detours the blocking central gate (CG) and forms a narrow channel through the transmembrane helices (TM) 2, 3, 6 and 7. In contrast, if E90 deprotonates after retinal isomerization, the primary proton acceptor is D253, and an almost-open channel through TM1, 2, 3 and 7 is generated. The channel diameter is very close to the experimental value. The potential mean force (PMF) suggests that the free energy is extremely low for ions passing through this channel.

Conventional Molecular Dynamics and Metadynamics Simulation Studies of the Binding and Unbinding Mechanism of TTR Stabilizers AG10 and Tafamidis.

Amyloid transthyretin (ATTR) amyloidosis is a widespread and fatal systemic amyloidosis characterized by the misfolding and amyloid aggregation of transthyretin (TTR). Studies suggest that dissociation of the TTR tetramer is the key step for its misfolding. Because of the importance of tetramer dissociation on ATTR amyloidosis, many TTR stabilizers have been discovered to stabilize the tetramer structure. This paper describes the application conventional molecular dynamics and metadynamics simulations to investigate the binding and unbinding mechanisms of two TTR stabilizers, including AG10 and tafamidis. AG10 has been granted an orphan drug designation by the U.S. Food and Drug Administration (FDA), and tafamidis was the first FDA-approved treatment for ATTR cardiomyopathy. The conventional molecular dynamics simulations reveal that both AG10 and tafamidis can stabilize the TTR tetramer through different mechanisms. AG10 stabilizes TTR tetramer by forming H-bonds with S117 to mimic the protective effect of T119M. Tafamidis stabilizes the tetramer by forming H-bond with S52 in the flexible CD loop to increase its structural stability. Despite the strong binding affinity of tafamidis, the free-energy surface constructed from metadynamics simulation suggests that tafamidis unbinds more readily than AG10 with lower free-energy barriers between the binding state and other intermediates. Finally, by performing pharmacophore analysis, we found two common important moieties of the studied compounds for their binding on the pockets, which can provide valuable guidance for future lead compounds' optimization in designing drugs for ATTR amyloidosis.

The role of Rho GTPases' substrates Rac and Cdc42 in osteoclastogenesis and relevant natural medicinal products study.

Recently, Rho GTPases substrates include Rac (Rac1 and Rac2) and Cdc42 that have been reported to exert multiple cellular functions in osteoclasts, the most prominent of which includes regulating the dynamic actin cytoskeleton rearrangements. In addition, natural products and their molecular frameworks have a long tradition as valuable starting points for medicinal chemistry and drug discovery. Although currently, there are reports about the natural product, which could play a therapeutic role in bone loss diseases (osteoporosis and osteolysis) through the regulation of Rac1/2 and Cdc42 during osteoclasts cytoskeletal structuring. There have been several excellent studies for exploring the therapeutic potentials of various natural products for their role in inhibiting cancer cells migration and function via regulating the Rac1/2 and Cdc42. Herein in this review, we try to focus on recent advancement studies for extensively understanding the role of Rho GTPases substrates Rac1, Rac2 and Cdc42 in osteoclastogenesis, as well as therapeutic potentials of natural medicinal products for their properties on the regulation of Rac1, and/or Rac2 and Cdc42, which is in order to inspire drug discovery in regulating osteoclastogenesis.

Molecular Dynamics Simulation of Transmembrane Transport of Chloride Ions in Mutants of Channelrhodopsin.

Channelrhodopsins (ChRs) are light-gated transmembrane cation channels which are widely used for optogenetic technology. Replacing glutamate located at the central gate of the ion channel with positively charged amino acid residues will reverse ion selectivity and allow anion conduction. The structures and properties of the ion channel, the transport of chloride, and potential of mean force (PMF) of the chimera protein (C1C2) and its mutants, EK-TC, ER-TC and iChloC, were investigated by molecular dynamics simulation. The results show that the five-fold mutation in E122Q-E129R-E140S-D195N-T198C (iChloC) increases the flexibility of the transmembrane channel protein better than the double mutations in EK-TC and ER-TC, and results in an expanded ion channel pore size and decreased steric resistance. The iChloC mutant was also found to have a higher affinity for chloride ions and, based on surface electrostatic potential analysis, provides a favorable electrostatic environment for anion conduction. The PMF free energy curves revealed that high affinity Cl- binding sites are generated near the central gate of the three mutant proteins. The energy barriers for the EK-TC and ER-TC were found to be much higher than that of iChloC. The results suggest that the transmembrane ion channel of iChloC protein is better at facilitating the capture and transport of chloride ions.

Exploration of the Misfolding Mechanism of Transthyretin Monomer: Insights from Hybrid-Resolution Simulations and Markov State Model Analysis.

Misfolding and aggregation of transthyretin (TTR) is widely known to be responsible for a progressive systemic disorder called amyloid transthyretin (ATTR) amyloidosis. Studies suggest that TTR aggregation is initiated by a rate-limiting dissociation of the homo-tetramer into its monomers, which can rapidly misfold and self-assemble into amyloid fibril. Thus, exploring conformational change involved in TTR monomer misfolding is of vital importance for understanding the pathogenesis of ATTR amyloidosis. In this work, microsecond timescale hybrid-resolution molecular dynamics (MD) simulations combined with Markov state model (MSM) analysis were performed to investigate the misfolding mechanism of the TTR monomer. The results indicate that a macrostate with partially unfolded conformations may serve as the misfolded state of the TTR monomer. This misfolded state was extremely stable with a very large equilibrium probability of about 85.28%. With secondary structure analysis, we found the DAGH sheet in this state to be significantly destroyed. The CBEF sheet was relatively stable and sheet structure was maintained. However, the F-strand in this sheet was likely to move away from E-strand and reform a new ß-sheet with the H-strand. This observation is consistent with experimental finding that F and H strands in the outer edge drive the misfolding of TTR. Finally, transition pathways from a near native state to this misfolded macrostate showed that the conformational transition can occur either through a native-like ß-sheet intermediates or through partially unfolded intermediates, while the later appears to be the main pathway. As a whole, we identified a potential misfolded state of the TTR monomer and elucidated the misfolding pathway for its conformational transition. This work can provide a valuable theoretical basis for understanding of TTR aggregation and the pathogenesis of ATTR amyloidosis at the atomic level.

Mesoscale Simulation to Study Constitutive Properties of TATB/F2314 PBX.

Material Point Method (MPM) mesoscale simulation was used to study the constitutive relation of a polymer bonded explosive (PBX) consisting of 1,3,5-triamino-2,4,6-trinitrobenzene (TATB) and a fluorine polymer binder F2314. The stress-strain variations of the PBX were calculated for different temperatures and different porosities, and the results were found to be consistent with experimental observations. The stress-strain relations at different temperatures were used to develop the constitutive equation of the PBX by using numerical data fitting. Stress-strain data for different porosities were used to establish the constitutive equation by fitting the simulation data to an improved Hashion-Shtrikman model. The equation can be used to predict the shear modulus and bulk modulus of the PBX at different densities of the sample. The constitutive equations developed for TATB/F2314 PBX by MPM mesoscale simulation are important equations for the numerical simulations of the PBX at macroscale. The method presented in this study provides an alternative approach for studying the constitutive relations of PBX.

Formation Mechanism of Ion Channel in Channelrhodopsin-2: Molecular Dynamics Simulation and Steering Molecular Dynamics Simulations.

Channelrhodopsin-2 (ChR2) is a light-activated and non-selective cationic channel protein that can be easily expressed in specific neurons to control neuronal activity by light. Although ChR2 has been extensively used as an optogenetic tool in neuroscience research, the molecular mechanism of cation channel formation following retinal photoisomerization in ChR2 is not well understood. In this paper, studies of the closed and opened state ChR2 structures are presented. The formation of the cationic channel is elucidated in atomic detail using molecular dynamics simulations on the all-trans-retinal (ChR2-trans) configuration of ChR2 and its isomerization products, 13-cis-retinal (ChR2-cis) configuration, respectively. Photoisomerization of the retinal-chromophore causes the destruction of interactions among the crucial residues (e.g., E90, E82, N258, and R268) around the channel and the extended H-bond network mediated by numerous water molecules, which opens the pore. Steering molecular dynamics (SMD) simulations show that the electrostatic interactions at the binding sites in intracellular gate (ICG) and central gate (CG) can influence the transmembrane transport of Na+ in ChR2-cis obviously. Potential of mean force (PMF) constructed by SMD and umbrella sampling also found the existing energy wells at these two binding sites during the transportation of Na+. These wells partly hinder the penetration of Na+ into cytoplasm through the ion channel. This investigation provides a theoretical insight on the formation mechanism of ion channels and the mechanism of ion permeation.

A Photophysical Deactivation Channel of Laser-Excited TATB Based on Semiclassical Dynamics Simulation and TD-DFT Calculation.

A deactivation channel for laser-excited 1,3,5-triamino-2,4,6-trinitrobenzene (TATB) was studied by semiclassical dynamics. Results indicate that the excited state resulting from an electronic transition from the highest occupied molecular orbital (HOMO) to the lowest unoccupied molecular mrbital (LUMO) is deactivated via pyramidalization of the activated N atom in a nitro group, with a lifetime of 2.4 ps. An approximately 0.5-electron transfer from the aromatic ring to the activated nitro group led to a significant increase of the C⁻NO2 bond length, which suggests that C⁻NO2 bond breaking could be a trigger for an explosive reaction. The time-dependent density functional theory (TD-DFT) method was used to calculate the energies of the ground and S1 excited states for each configuration in the simulated trajectory. The S1←S0 energy gap at the instance of non-adiabatic decay was found to be 0.096 eV, suggesting that the decay geometry is close to the conical intersection.

Thymine dimer splitting in the T<>T-G trinucleotide model system: a semiclassical dynamics and TD-DFT study.

The mechanism leading to bond cleavage of a thymine-thymine cyclobutane dimer (T<>T) in a model system consisting of the dimer flanked by guanine trinucleotide was studied using semiclassical dynamics simulation. Pulsed laser excitation of the guanine molecule is found to cause electron transfer from the guanine molecule to the dimer, which then dissociates via sequential cleavage of the C5C5' and C6C6' bonds. Subsequently, electrons transfer back to the guanine molecule as the dimer splits into two monomers. The splitting of the cyclobutane dimer was found to be in the femtosecond time scale.

Dynamics of laser-excited stacked adenines: semiclassical simulations.

The nonradiative decay of a π-stacked pair of adenine molecules, following laser excitation, was studied by semiclassical dynamics simulations. Two deactivation pathways were characterized. One pathway involves an ultrafast internal conversion within ~600 fs induced by an out-of-plane vibration of the H atom and deformation of the pyrimidine ring at the C(2) site. A slower process (~2400 fs) involves covalent bond formation between the stacked molecules, which lowers the excimer state energy and inhibits the deformation of the pyrimidine ring; the decay is also induced by an out-of-plane vibration of the H atom at the C(2) site of the pyrimidine ring.

Detailed mechanism for photoinduced cytosine dimerization: a semiclassical dynamics simulation.

Semiclassical dynamics simulation is used to study dimerization of two stacked cytosine molecules following excitation by ultrashort laser pulses (25 fs fwhm, Gaussian, 4.1 eV photon energy). The initial excited state was found to form an ultrashort exciton state, which eventually leads to the formation of an excimer state by charge transfer. When the interbase distance, defined as an average value of C(5)-C(5)' and C(6)-C(6)', becomes less than 3 Å, charge recombination occurs due to strong intermolecular interaction, eventually leading to an avoided crossing within 20-30 fs. Geometries at the avoided crossing, with average intermolecular distance of about 2.1 Å, are in accord with CASSCF/CASPT2 calculations. Results indicate that the C(2)-N(1)-C(6)-C(5) and C(2)'-N(1)'-C(6)'-C(5)' dihedral angles' bending vibrations play a significant role in the vibronic coupling between the HOMO and LUMO, which leads to a nonadiabatic transition to the electronic ground state.

Laser induced C(60) cage opening studied by semiclassical dynamics simulation.

Laser induced opening of the C(60) cage is studied by a semiclassical electron-radiation-ion dynamics technique. The simulation results indicate that the C(60) cage is abruptly opened immediately after laser excitation. The opening of the C(60) cage induces a quick increase in kinetic energy and a sharp decrease in electronic energy, suggesting that the breaking of the C(60) cage efficiently heats up the cluster and enhances the thermal fragmentation of C(60) fullerene.

Photoinduced dissociation of cyclobutane thymine dimer studied by semiclassical dynamics simulation.

Photoinduced dissociation of the thymine dimer is studied in a semiclassical dynamics simulation. The simulation follows excitation of an isolated thymine dimer by a 25 fs fwhm laser pulse, and finds that dissociation proceeds via an asynchronously concerted mechanism, in which the C(5)-C(5)' bond breaks soon after application of the laser pulse, followed by cleavage of the C(6)-C(6)' bond. The dissociation results in two thymine monomers, one in an electronically excited state and the other in the ground state. The former decays to the electronic ground state through an avoided crossing induced by deformation of the pyrimidine ring at the C(5)' and C(6)' sites.

New implementations of MRCI in semiempirical frameworks.

Multireference configuration interaction with single and double excitations (MRCISD) as well as its analytic CI gradients has been implemented in the semiempirical framework. The hole-particle symmetry and a mixed driven model for computing coupling coefficients have been used in the new code that allows us to perform MRCI and gradient calculations with higher efficiency and less storage requirements.

Nonadiabatic simulation study of photoisomerization of azobenzene: detailed mechanism and load-resisting capacity.

Nonadiabatic dynamical simulations were carried out to study cis-to-trans isomerization of azobenzene under laser irradiation and/or external mechanical loads. We used a semiclassical electron-radiation-ion dynamics method that is able to describe the coevolution of the structural dynamics and the underlying electronic dynamics in a real-time manner. It is found that azobenzene photoisomerization occurs predominantly by an out-of-plane rotation mechanism even under a nontrivial resisting force of several tens of piconewtons. We have repeated the simulations systematically for a broad range of parameters for laser pulses, but could not find any photoisomerization event by a previously suggested in-plane inversion mechanism. The simulations found that the photoisomerization process can be held back by an external resisting force of 90-200 pN depending on the frequency and intensity of the lasers. This study also found that a pure mechanical isomerization is possible from the cis-to-trans state if the azobenzene molecule is stretched by an external force of approximately 1250-1650 pN. Remarkably, the mechanical isomerization first proceeds through a mechanically activated inversion, and then is diverted to an ultrafast downhill rotation that accomplishes the isomerization. Implications of these findings to azobenzene-based nanomechanical devices are discussed.

Why does trans-azobenzene have a smaller isomerization yield for pi pi* excitation than for n pi* excitation?

A realistic dynamics simulation study is reported for the photoisomerization of trans-azobenzene. The isomerization follows both n pi* (the HOMO --> LUMO) and pi pi* (the HOMO-1 --> LUMO) excitations. The simulation finds that for the pi pi* excitation, the relaxation of the S(pi pi*) state is immediately followed by double excitation, (pi)(2)(pi*)(2). The decay from the S((pi)(2)(pi*)(2)) state to the S(0) state can occur at partially twisted structure, which favors the formation of the trans isomer. Multiple decay channels are found at about twisted structure for both n pi* and pi pi* excitations. Decay at about twisted geometry leads to the formation of either cis or trans isomer. Opening of the decay channel at partially twisted structure accounts for the smaller isomerization yield for the pi pi* excitation.

Detailed dynamics of the nonradiative deactivation of adenine: a semiclassical dynamics study.

A realistic dynamics simulation study is reported for the ultrafast radiationless deactivation of 9H-adenine. The simulation follows two different excitations induced by two 80 fs (fwhm) laser pulses that are different in energy: one has a photon energy of 5.0 eV, and the other has a photon energy of 4.8 eV. The simulation shows that the excited molecule decays to the electronic ground state from the (1)pipi* state in both excitations but through two different radiationless pathways: in the 5.0 eV excitation, the decay channel involves the out-of-plane vibration of the amino group, whereas in the 4.8 eV excitation, the decay strongly associates with the deformation of the pyrimidine at the C 2 atom. The lifetime of the (1) npi* state determined in the simulation study is 630 fs for the 5.0 eV excitation and 1120 fs for the 4.8 eV excitation. These are consistent with the experimental values of 750 and 1000 fs. We conclude that the experimentally observed difference in the lifetime of the (1) npi* state at various excitations results from the different radiationless deactivation pathways of the excited molecule to the electronic ground state.