RISMiCal: A software package to perform fast RISM/3D-RISM calculations.

Solvent plays an essential role in a variety of chemical, physical, and biological processes that occur in the solution phase. The reference interaction site model (RISM) and its three-dimensional extension (3D-RISM) serve as powerful computational tools for modeling solvation effects in chemical reactions, biological functions, and structure formations. We present the RISM integrated calculator (RISMiCal) program package, which is based on RISM and 3D-RISM theories with fast GPU code. RISMiCal has been developed as an integrated RISM/3D-RISM program that has interfaces with external programs such as Gaussian16, GAMESS, and Tinker. Fast 3D-RISM programs for single- and multi-GPU codes written in CUDA would enhance the availability of these hybrid methods because they require the performance of many computationally expensive 3D-RISM calculations. We expect that our package can be widely applied for chemical and biological processes in solvent. The RISMiCal package is available at https://rismical-dev.github.io.

Analysis of Protein Folding Simulation with Moving Root Mean Square Deviation.

We apply moving root-mean-square deviation (mRMSD), which does not require a reference structure, as a method for analyzing protein dynamics. This method can be used to calculate the root-mean-square deviation (RMSD) of structure between two specified time points and to analyze protein dynamics behavior through time series analysis. We applied this method to the Trp-cage trajectory calculated by the Anton supercomputer and found that it shows regions of stable states as well as the conventional RMSD. In addition, we extracted a characteristic structure in which the side chains of Asp1 and Arg16 form hydrogen bonds near the most stable structure of the Trp-cage. We also determined that ≥20 ns is an appropriate time interval to investigate protein dynamics using mRMSD. Applying this method to NuG2 protein, we found that mRMSD can be used to detect regions of metastable states in addition to the stable state. This method can be applied to molecular dynamics simulations of proteins whose stable structures are unknown.

Volatile Short-Chain Aliphatic Aldehydes Act as Taste Modulators through the Orally Expressed Calcium-Sensing Receptor CaSR.

Aldehydes are natural volatile aroma compounds generated by the Maillard reaction of sugars and amino acids in food and affect the flavor of food. They have been reported to exert taste-modifying effects, such as increases in taste intensity at concentrations below the odor detection threshold. The present study examined the taste-enhancing effects of short-chain aliphatic aldehydes, such as isovaleraldehyde (IVAH) and 2-methylbutyraldehyde, thus attempting to identify the taste receptors involved. The results obtained revealed that IVAH enhanced the taste intensity of taste solutions even under the condition of olfactory deprivation by a noseclip. Furthermore, IVAH activated the calcium-sensing receptor CaSR in vitro. Receptor assays on aldehyde analogues showed that C3-C6 aliphatic aldehydes and methional, a C4 sulfur aldehyde, activated CaSR. These aldehydes functioned as a positive allosteric modulator for CaSR. The relationship between the activation of CaSR and taste-modifying effects was investigated by a sensory evaluation. Taste-modifying effects were found to be dependent on the activation state of CaSR. Collectively, these results suggest that short-chain aliphatic aldehydes function as taste modulators that modify sensations by activating orally expressed CaSR. We propose that volatile aroma aldehydes may also partially contribute to the taste-modifying effect via the same molecular mechanism as kokumi substances.

Computational Analysis of the SARS-CoV-2 RBD-ACE2-Binding Process Based on MD and the 3D-RISM Theory.

The binding process of angiotensin-converting enzyme 2 (ACE2) to the receptor-binding domain (RBD) of the severe acute respiratory syndrome-like coronavirus 2 spike protein was investigated using molecular dynamics simulation and the three-dimensional reference interaction-site model theory. The results suggested that the protein-binding process consists of a protein-protein approaching step, followed by a local structural rearrangement step. In the approaching step, the interprotein interaction energy decreased as the proteins approached each other, whereas the solvation free energy increased. As the proteins approached, the glycan of ACE2 first established a hydrogen bond with the RBD. Thereafter, the number of interprotein hydrogen bonds increased rapidly. The solvation free energy increased because of the desolvation of the protein as it approached its partner. The spatial distribution function of the solvent revealed the presence of hydrogen bonds bridged by water molecules on the RBD-ACE2 interface. Finally, principal component analysis revealed that ACE2 showed a pronounced conformational change, whereas there was no significant change in RBD.

Application of reference-modified density functional theory: Temperature and pressure dependences of solvation free energy.

Recently, we proposed a reference-modified density functional theory (RMDFT) to calculate solvation free energy (SFE), in which a hard-sphere fluid was introduced as the reference system instead of an ideal molecular gas. Through the RMDFT, using an optimal diameter for the hard-sphere reference system, the values of the SFE calculated at room temperature and normal pressure were in good agreement with those for more than 500 small organic molecules in water as determined by experiments. In this study, we present an application of the RMDFT for calculating the temperature and pressure dependences of the SFE for solute molecules in water. We demonstrate that the RMDFT has high predictive ability for the temperature and pressure dependences of the SFE for small solute molecules in water when the optimal reference hard-sphere diameter determined for each thermodynamic condition is used. We also apply the RMDFT to investigate the temperature and pressure dependences of the thermodynamic stability of an artificial small protein, chignolin, and discuss the mechanism of high-temperature and high-pressure unfolding of the protein. © 2017 Wiley Periodicals, Inc.

A reference-modified density functional theory: An application to solvation free-energy calculations for a Lennard-Jones solution.

In the conventional classical density functional theory (DFT) for simple fluids, an ideal gas is usually chosen as the reference system because there is a one-to-one correspondence between the external field and the density distribution function, and the exact intrinsic free-energy functional is available for the ideal gas. In this case, the second-order density functional Taylor series expansion of the excess intrinsic free-energy functional provides the hypernetted-chain (HNC) approximation. Recently, it has been shown that the HNC approximation significantly overestimates the solvation free energy (SFE) for an infinitely dilute Lennard-Jones (LJ) solution, especially when the solute particles are several times larger than the solvent particles [T. Miyata and J. Thapa, Chem. Phys. Lett. 604, 122 (2014)]. In the present study, we propose a reference-modified density functional theory as a systematic approach to improve the SFE functional as well as the pair distribution functions. The second-order density functional Taylor series expansion for the excess part of the intrinsic free-energy functional in which a hard-sphere fluid is introduced as the reference system instead of an ideal gas is applied to the LJ pure and infinitely dilute solution systems and is proved to remarkably improve the drawbacks of the HNC approximation. Furthermore, the third-order density functional expansion approximation in which a factorization approximation is applied to the triplet direct correlation function is examined for the LJ systems. We also show that the third-order contribution can yield further refinements for both the pair distribution function and the excess chemical potential for the pure LJ liquids.

Structure-CaSR-Activity Relation of Kokumi γ-Glutamyl Peptides.

Modulation of the calcium sensing receptor (CaSR) is one of the physiological activities of γ-glutamyl peptides such as glutathione (γ-glutamylcysteinylglycine). γ-Glutamyl peptides also possess a flavoring effect, i.e., sensory activity of kokumi substances, which modifies the five basic tastes when added to food. These activities have been shown to be positively correlated, suggesting that kokumi γ-glutamyl peptides are perceived through CaSRs in humans. Our research is based on the hypothesis that the discovery of highly active CaSR agonist peptides will lead to the creation of practical kokumi peptides. Through continuous study of the structure-CaSR-activity relation of a large number of γ-glutamyl peptides, we have determined that the structural requirements for intense CaSR activity of γ-glutamyl peptides are as follows: existence of an N-terminal γ-L-glutamyl residue; existence of a moderately sized, aliphatic, neutral substituent at the second residue in an L-configuration; and existence of a C-terminal carboxylic acid, preferably with the existence of glycine as the third constituent. By the sensory analysis of γ-glutamyl peptides selected by screening using the CaSR activity assay, γ-glutamylvalylglycine was found to be a potent kokumi peptide. Furthermore, norvaline-containing γ-glutamyl peptides, i.e., γ-glutamylnorvalylglycine and γ-glutamylnorvaline, possessed excellent sensory activity of kokumi substances. A novel, practical industrial synthesis of regiospecific γ-glutamyl peptides is also required for their commercialization, which was achieved through the ring opening reaction of N-α-carbobenzoxy-L-glutamic anhydride and amino acids or peptides in the presence of N-hydroxysuccinimide.

A solvation-free-energy functional: a reference-modified density functional formulation.

The three-dimensional reference interaction site model (3D-RISM) theory, which is one of the most applicable integral equation theories for molecular liquids, overestimates the absolute values of solvation-free-energy (SFE) for large solute molecules in water. To improve the free-energy density functional for the SFE of solute molecules, we propose a reference-modified density functional theory (RMDFT) that is a general theoretical approach to construct the free-energy density functional systematically. In the RMDFT formulation, hard-sphere (HS) fluids are introduced as the reference system instead of an ideal polyatomic molecular gas, which has been regarded as the appropriate reference system of the interaction-site-model density functional theory for polyatomic molecular fluids. We show that using RMDFT with a reference HS system can significantly improve the absolute values of the SFE for a set of neutral amino acid side-chain analogues as well as for 504 small organic molecules.

Mesophyll conductance in leaves of Japanese white birch (Betula platyphylla var. japonica) seedlings grown under elevated CO2 concentration and low N availability.

To test the hypothesis that mesophyll conductance (gm ) would be reduced by leaf starch accumulation in plants grown under elevated CO2 concentration [CO2 ], we investigated gm in seedlings of Japanese white birch grown under ambient and elevated [CO2 ] with an adequate and limited nitrogen supply using simultaneous gas exchange and chlorophyll fluorescence measurements. Both elevated [CO2 ] and limited nitrogen supply decreased area-based leaf N accompanied with a decrease in the maximum rate of Rubisco carboxylation (Vc,max ) on a CO2 concentration at chloroplast stroma (Cc ) basis. Conversely, only seedlings grown at elevated [CO2 ] under limited nitrogen supply had significantly higher leaf starch content with significantly lower gm among the treatment combinations. Based on a leaf anatomical analysis using microscopic photographs, however, there were no significant difference in the area of chloroplast surfaces facing intercellular space per unit leaf area among treatment combinations. Thicker cell walls were suggested in plants grown under limited N by increases in leaf mass per area subtracting non-structural carbohydrates. These results suggest that starch accumulation and/or thicker cell walls in the leaves grown at elevated [CO2 ] under limited N supply might hinder CO2 diffusion in chloroplasts and cell walls, which would be an additional cause of photosynthetic downregulation as well as a reduction in Rubisco activity related to the reduced leaf N under elevated [CO2 ].

Massively parallel implementation of 3D-RISM calculation with volumetric 3D-FFT.

A new three-dimensional reference interaction site model (3D-RISM) program for massively parallel machines combined with the volumetric 3D fast Fourier transform (3D-FFT) was developed, and tested on the RIKEN K supercomputer. The ordinary parallel 3D-RISM program has a limitation on the number of parallelizations because of the limitations of the slab-type 3D-FFT. The volumetric 3D-FFT relieves this limitation drastically. We tested the 3D-RISM calculation on the large and fine calculation cell (2048(3) grid points) on 16,384 nodes, each having eight CPU cores. The new 3D-RISM program achieved excellent scalability to the parallelization, running on the RIKEN K supercomputer. As a benchmark application, we employed the program, combined with molecular dynamics simulation, to analyze the oligomerization process of chymotrypsin Inhibitor 2 mutant. The results demonstrate that the massive parallel 3D-RISM program is effective to analyze the hydration properties of the large biomolecular systems.

Effect of Main and Side Chains on the Folding Mechanism of the Trp-Cage Miniprotein.

Proteins that do not fold into their functional native state have been linked to diseases. In this study, the influence of the main and side chains of individual amino acids on the folding of the tryptophan cage (Trp-cage), a designed 20-residue miniprotein, was analyzed. For this purpose, we calculated the solvation free energy (SFE) contributions of individual atoms by using the 3D-reference interaction site model with the atomic decomposition method. The mechanism by which the Trp-cage is stabilized during the folding process was examined by calculating the total energy, which is the sum of the conformational energy and SFE. The folding process of the Trp-cage resulted in a stable native state, with a total energy that was 62.4 kcal/mol lower than that of the unfolded state. The solvation entropy, which is considered to be responsible for the hydrophobic effect, contributed 31.3 kcal/mol to structural stabilization. In other words, the contribution of the solvation entropy accounted for approximately half of the total contribution to Trp-cage folding. The hydrophobic core centered on Trp6 contributed 15.6 kcal/mol to the total energy, whereas the solvation entropy contribution was 6.3 kcal/mol. The salt bridge formed by the hydrophilic side chains of Asp9 and Arg16 contributed 10.9 and 5.0 kcal/mol, respectively. This indicates that not only the hydrophobic core but also the salt bridge of the hydrophilic side chains gain solvation entropy and contribute to stabilizing the native structure of the Trp-cage.

Leaves of Japanese oak (Quercus mongolica var. crispula) mitigate photoinhibition by adjusting electron transport capacities and thermal energy dissipation along the intra-canopy light gradient.

We investigated the morphological and physiological acclimation of leaves grown within a canopy of Japanese oak tree (Quercus mongolica var. crispula) in terms of the susceptibility to photoinhibition under various growth light conditions. The maximum rates of photosynthesis (P(max)) and electron transport (ETR(max)) were higher in mature leaves grown under stronger light with higher area-based leaf nitrogen (N) content closely associated with higher leaf mass per area. The net photosynthetic (P(n)) and electron transport (ETR) rates corresponding to the daily peak photosynthetic photon flux density (PPFD(max)) during leaf maturation were almost comparable to P(max) and ETR(max), respectively. Conversely, P(n) and ETR at the daily average PPFD (PPFD(avg)) were substantially low in shade-grown leaves when compared with P(max) and ETR(max). The susceptibility to photoinhibition at PPFD(max), i.e. at sunflecks for the shade-grown leaves, was assessed by the rate of excess energy production. Although sun leaves showed higher rates of electron transport and thermal energy dissipation than shade leaves under PPFD(max) conditions, the rate of excess energy production was almost constant across shade to sun leaves. The shade leaves of the Japanese oak grown within a crown were suggested to adjust their N investment to maintain higher photosynthetic capacities compared with those required to maximize the net carbon gain, which may facilitate the dissipation of the excessive light energy of sunflecks to circumvent photoinhibition in cooperation with thermal energy dissipation.

Increases in the pungency of allyl isothiocyanate and piperine by CaSR agonists, glutathione and γ-glutamyl-valyl-glycine.

γ-Glutamyl peptides, including glutathione (γ-Glu-Cys-Gly, GSH) and γ-glutamyl-valyl-glycine (γ-Glu-Val-Gly), have been shown to increase the intensity of basic tastes, such as salty, sweet, and umami, and flavor, including mouthfulness, but had no taste themselves at the concentrations tested. Although the mechanisms of action of γ-glutamyl peptides currently remain unclear, the involvement of the calcium sensing receptor (CaSR) has been suggested. Since GSH and γ-Glu-Val-Gly increase the pungency of some spices, the present study investigated their effects on the pungency of allyl isothiocyanate (AITC) using a sensory evaluation. GSH and γ-Glu-Val-Gly both significantly increased the pungency of AITC, while anserine, a peptide without CaSR activity, did not. GSH-induced increases in pungency were suppressed by NPS-2143, a CaSR inhibitor. Further, γ-Glu-Val-Gly significantly increased the pungency of piperine. The present results suggest that GSH and γ-Glu-Val-Gly increased the pungency by activating CaSR.

Lipocalin 15 in the olfactory mucus is a biomarker for Bowman's gland activity.

Olfactory mucus contributes to the specific functions of the olfactory mucosa, but the composition and source of mucus proteins have not been fully elucidated. In this study, we used comprehensive proteome analysis and identified lipocalin 15 (LCN15), a human-specific lipocalin family protein, as an abundant component of the olfactory mucus. Western blot analysis and enzyme-linked immunosorbent assay (ELISA) using a newly generated anti-LCN15 antibody showed that LCN15 was concentrated in olfactory mucus samples, but not in respiratory mucus samples. Immunohistochemical staining using anti-LCN15 antibody revealed that LCN15 localized to the cytokeratin 18-positive Bowman's glands of the olfactory cleft mucosa. Quantitative image analysis revealed that the area of LCN15 immunoreactivity along the olfactory cleft mucosa significantly correlated with the area of neuron-specific Protein-Gene Product 9.5 (PGP9.5) immunoreactivity, suggesting that LCN15 is produced in non-degenerated areas of the olfactory neuroepithelium. ELISA demonstrated that the concentration of LCN15 in the mucus was lower in participants with normal olfaction (≥ 50 years) and also tended to be lower in patients with idiopathic olfactory loss (≥ 50 years) than in participants with normal olfaction (< 50 years). Thus, LCN15 may serve as a biomarker for the activity of the Bowman's glands.

Microplastic pollution of commercial fishes from coastal and offshore waters in southwestern Japan.

Microplastic (MP) pollution in the marine environment is a worldwide issue. There is growing concern of consuming MPs through fish, yet the contamination status of fish collected from deeper waters surrounding Japan remains limited. Here, we presented baseline data on MPs in commercially important fishes from the coastal and offshore waters near Kyushu, Japan (East China Sea). We examined the MPs in the digestive tracts of two pelagic (n = 150) and five demersal species (n = 235). The fish were caught by pole and line, and bottom trawl at different geographical positions. The MPs in pelagic fish (39.1%) were more than in demersal fish (10.3%) and were of larger sizes. Moreover, the MPs correlated with habitat depth and type and species variation in the shape and polymer composition of MPs was observed. The results increase our understanding of the heterogeneous uptake of MPs by fishes.

Involvement of the calcium-sensing receptor in human taste perception.

By human sensory analyses, we found that various extracellular calcium-sensing receptor (CaSR) agonists enhance sweet, salty, and umami tastes, although they have no taste themselves. These characteristics are known as "kokumi taste" and often appear in traditional Japanese cuisine. Although GSH is a typical kokumi taste substance (taste enhancer), its mode of action is poorly understood. Here, we demonstrate how the kokumi taste is enhanced by the CaSR, a close relative of the class C G-protein-coupled receptors T1R1, T1R2, and T1R3 (sweet and umami receptors). We identified a large number of CaSR agonist gamma-glutamyl peptides, including GSH (gamma-Glu-Cys-Gly) and gamma-Glu-Val-Gly, and showed that these peptides elicit the kokumi taste. Further analyses revealed that some known CaSR agonists such as Ca(2+), protamine, polylysine, L-histidine, and cinacalcet (a calcium-mimetic drug) also elicit the kokumi taste and that the CaSR-specific antagonist, NPS-2143, significantly suppresses the kokumi taste. This is the first report indicating a distinct function of the CaSR in human taste perception.

Activation of polycystic kidney disease-2-like 1 (PKD2L1)-PKD1L3 complex by acid in mouse taste cells.

Five basic tastes (bitter, sweet, umami, salty, and sour) are detected in the four taste areas where taste buds reside. Although molecular mechanisms for detecting bitter, sweet, and umami have been well clarified, those for sour and salty remain poorly understood. Several channels including acid-sensing ion channels have been proposed as candidate sour receptors, but they do not encompass all sour-sensing abilities in vivo. We recently reported a novel candidate for sour sensing, the polycystic kidney disease-2-like 1 (PKD2L1)-PKD1L3 channel complex. This channel is not a traditional ligand-gated channel and is gated open only after removal of an acid stimulus, called an off response. Here we show that off responses upon acid stimulus are clearly observed in native taste cells from circumvallate, but not fungiform papillae, of glutamate decarboxylase 67-green fluorescent protein (GAD67-GFP) knock-in mice, from which Type III taste cells can be visualized, using Ca(2+) imaging and patch clamp methods. Off responses were detected in most cells where PKD2L1 immunoreactivity was observed. Interestingly, the pH threshold for acid-evoked intracellular Ca(2+) increase was around 5.0, a value much higher than that observed in HEK293 cells expressing the PKD2L1-PKD1L3 complex. Thus, PKD2L1-PKD1L3-mediated acid-evoked off responses occurred both in HEK293 cells and in native taste cells, suggesting the involvement of the PKD2L1-PKD1L3 complex in acid sensing in vivo.

GPR120 agonists enhance the fatty orosensation when added to fat-containing system, but do not evoke it by themselves in humans.

Dietary fat, an important macronutrient, has been considered to be perceived by texture and olfaction. Recently, fatty acid transporter, CD36, and fatty acid receptor, GPR120 are considered to be involved in human gustatory fatty acids perception in humans. However, limited information is currently available to show that agonists of CD36 and GPR120 evoke fatty oral sensations regarding to dietary fat in humans. Therefore, the role of GPR120 agonists in dietary fat perception in humans was investigated herein. An emulsion prepared from vegetable oil had a stronger fatty orosensation, an orosensation similar to an oily mouth-coating sensed 5 - 10 s after tasting, than that prepared from mineral oil; however, the physical properties of both emulsions, such as viscosity, particle distribution, interfacial tension, contact angle, frictional load, and ζ-electric potential were similar. The potent GPR120 agonist, TUG-891 enhanced the fatty orosensation when added to the emulsion prepared from vegetable oil, but not to that from mineral oil. All GPR120 agonists tested enhanced the fatty orosensation when added to a low-fat food system whereas they did not evoke any fatty sensation in aqueous solution at the concentrations tested in food system, and sensory activity positively correlated with GPR120 activity. These results suggest that GPR120 agonists enhance the fatty orosensation in humans when added to vegetable oil or a low-fat food system, but do not evoke it by themselves.

Random walk to describe diffusion phenomena in three-dimensional discontinuous media: Extension to anisotropic phases and nonplane interfaces.

We extend the step-balanced random walk [Y. Maruyama, Phys. Rev. E 96, 032135 (2017)2470-004510.1103/PhysRevE.96.032135], which was proposed for diffusion phenomena in three-dimensional discontinuous media, to systems that contain anisotropic phase zones with nonplane interfaces. What is key is threefold: For each interstep transition at discontinuous interfaces to be equilibrated by its reverse transition, incident and penetration steps are in one-to-one correspondence; at each incidence, penetration probability is determined by the normal components of an incident step and the corresponding penetration step with respect to an incident tangential plane, and for reflection, the reverse of an incident step, which satisfies the conditions for time reversibility at any interface, is used as a reflection step.