Development of equation of motion deciphering locomotion including omega turns of Caenorhabditis elegans.

Locomotion is a fundamental behavior of Caenorhabditis elegans (C. elegans). Previous works on kinetic simulations of animals helped researchers understand the physical mechanisms of locomotion and the muscle-controlling principles of neuronal circuits as an actuator part. It has yet to be understood how C. elegans utilizes the frictional forces caused by the tension of its muscles to perform sequenced locomotive behaviors. Here, we present a two-dimensional rigid body chain model for the locomotion of C. elegans by developing Newtonian equations of motion for each body segment of C. elegans. Having accounted for friction-coefficients of the surrounding environment, elastic constants of C. elegans, and its kymogram from experiments, our kinetic model (ElegansBot) reproduced various locomotion of C. elegans such as, but not limited to, forward-backward-(omega turn)-forward locomotion constituting escaping behavior and delta-turn navigation. Additionally, ElegansBot precisely quantified the forces acting on each body segment of C. elegans to allow investigation of the force distribution. This model will facilitate our understanding of the detailed mechanism of various locomotive behaviors at any given friction-coefficients of the surrounding environment. Furthermore, as the model ensures the performance of realistic behavior, it can be used to research actuator-controller interaction between muscles and neuronal circuits.

Development of quantitative and continuous measure for severity degree of Alzheimer's disease evaluated from MRI images of 761 human brains.

BACKGROUND: Alzheimer's disease affects profoundly the quality of human behavior and cognition. The very broad distribution of its severity across various human subjects requires the quantitative diagnose of Alzheimer's disease beyond the conventional tripartite classification of cohorts such as cognitively normal (CN), mild cognitive impairment (MCI), Alzheimer's disease (AD). The unfolding of such broad distributions by the quantitative and continuous degree of AD severity is necessary for the precise diagnose in the cross-sectional study of different stages in AD. RESULTS: We conducted the massive reanalysis on MRI images of 761 human brains based on the accumulated bigdata of Alzheimer's Disease Neuroimaging Initiative. The score matrix of cortical thickness profile at cortex points of subjects was constructed by statistically learning the cortical thickness data of 761 human brains. We also developed a new and simple algebraic predictor which provides the quantitative and continuous degree of AD severity of subjects along the scale from 0 for fully CN to 1 for fully AD state. The mathematical measure of a new predictor for the degree of AD severity is presented based on a covariance correlation matrix of cortical thickness profile between human subjects. One can remove the uncertainty in the determination of different stages in AD by the quantitative degree of AD severity and thus go far beyond the tripartite classification of cohorts. CONCLUSIONS: We unfold the nature of broad distribution of AD severity of subjects even within a given cohort by the scale from 0 for fully CN to 1 for fully AD state. The quantitative and continuous degree of AD severity developed in this study would be a good practical measure for diagnosing the different stages in AD severity.

Computational design of a thermolabile uracil-DNA glycosylase of Escherichia coli.

Polymerase chain reaction (PCR) is a powerful tool to diagnose infectious diseases. Uracil DNA glycosylase (UDG) is broadly used to remove carryover contamination in PCR. However, UDG can contribute to false negative results when not inactivated completely, leading to DNA degradation during the amplification step. In this study, we designed novel thermolabile UDG derivatives by supercomputing molecular dynamic simulations and residual network analysis. Based on enzyme activity analysis, thermolability, thermal stability, and biochemical experiments of Escherichia coli-derived UDG and 22 derivatives, we uncovered that the UDG D43A mutant eliminated the false negative problem, demonstrated high efficiency, and offered great benefit for use in PCR diagnosis. We further obtained structural and thermodynamic insights into the role of the D43A mutation, including perturbed protein structure near D43; weakened pairwise interactions of D43 with K42, N46, and R80; and decreased melting temperature and native fraction of the UDG D43A mutant compared with wild-type UDG.

Structural and molecular insight into the pH-induced low-permeability of the voltage-gated potassium channel Kv1.2 through dewetting of the water cavity.

Understanding the gating mechanism of ion channel proteins is key to understanding the regulation of cell signaling through these channels. Channel opening and closing are regulated by diverse environmental factors that include temperature, electrical voltage across the channel, and proton concentration. Low permeability in voltage-gated potassium ion channels (Kv) is intimately correlated with the prolonged action potential duration observed in many acidosis diseases. The Kv channels consist of voltage-sensing domains (S1-S4 helices) and central pore domains (S5-S6 helices) that include a selectivity filter and water-filled cavity. The voltage-sensing domain is responsible for the voltage-gating of Kv channels. While the low permeability of Kv channels to potassium ion is highly correlated with the cellular proton concentration, it is unclear how an intracellular acidic condition drives their closure, which may indicate an additional pH-dependent gating mechanism of the Kv family. Here, we show that two residues E327 and H418 in the proximity of the water cavity of Kv1.2 play crucial roles as a pH switch. In addition, we present a structural and molecular concept of the pH-dependent gating of Kv1.2 in atomic detail, showing that the protonation of E327 and H418 disrupts the electrostatic balance around the S6 helices, which leads to a straightening transition in the shape of their axes and causes dewetting of the water-filled cavity and closure of the channel. Our work offers a conceptual advancement to the regulation of the pH-dependent gating of various voltage-gated ion channels and their related biological functions.

The C-Domain of the NAC Transcription Factor ANAC019 Is Necessary for pH-Tuned DNA Binding through a Histidine Switch in the N-Domain.

The affinity of transcription factors (TFs) for their target DNA is a critical determinant of gene expression. Whether the DNA-binding domain (DBD) of TFs alone can regulate binding affinity to DNA is an important question for identifying the design principle of TFs. We studied ANAC019, a member of the NAC TF family of proteins in Arabidopsis, and found a well-conserved histidine switch located in its DBD, which regulates both homodimerization and transcriptional control of the TF through H135 protonation. We found that the removal of a C-terminal intrinsically disordered region (IDR) in the TF abolished the pH-dependent binding of the N-terminal DBD to DNA. We propose a mechanism in which long-range electrostatic interactions between DNA and the negatively charged C-terminal IDR turns on the pH dependency of the DNA-binding affinity of the N-terminal DBD.

Design of Reduction Process of SnO2 by CH4 for Efficient Sn Recovery.

We design a novel method for the CH4 reduction of SnO2 for the efficient recovery of Sn from SnO2 through a study combining theory and experiment. The atomic-level process of CH4-SnO2 interaction and temperature-dependent reduction behavior of SnO2 were studied with a combination of a multi-scale computational method of thermodynamic simulations and density functional theory (DFT) calculations. We found that CH4 was a highly efficient and a versatile reducing agent, as the total reducing power of CH4 originates from the carbon and hydrogen of CH4, which sequentially reduce SnO2. Moreover, as a result of the CH4 reduction of SnO2, a mixture of CO and H2 was produced as a gas-phase product (syngas). The relative molar ratio of the produced gas-phase product was controllable by the reduction temperature and the amount of supplied CH4. The laboratory-scale experimental study confirmed that CH4 actively reduces SnO2, producing 99.34% high-purity Sn and H2 and CO. Our results present a novel method for an efficient, green, and economical recycling strategy for Sn with economic value added that is held by the co-produced clean energy source (syngas).

Effect of Korean Red Ginseng in chronic liver disease.

Chronic liver disease, one of the most common diseases, typically arises from nonalcoholic fatty liver disease, alcoholic liver disease, chronic viral hepatitis, or hepatocellular carcinoma. Therefore, there is a pressing need for improved treatment strategies. Korean Red Ginseng has been known to have positive effects on liver disease and liver function. In this paper, we summarize the current knowledge on the beneficial effects of Korean Red Ginseng on chronic liver disease, a condition encompassing nonalcoholic fatty liver disease, alcoholic liver disease, chronic viral hepatitis, and hepatocellular carcinoma, as supported by experimental evaluation and clinical investigation.

Probing the Folding-Unfolding Transition of a Thermophilic Protein, MTH1880.

The folding mechanism of typical proteins has been studied widely, while our understanding of the origin of the high stability of thermophilic proteins is still elusive. Of particular interest is how an atypical thermophilic protein with a novel fold maintains its structure and stability under extreme conditions. Folding-unfolding transitions of MTH1880, a thermophilic protein from Methanobacterium thermoautotrophicum, induced by heat, urea, and GdnHCl, were investigated using spectroscopic techniques including circular dichorism, fluorescence, NMR combined with molecular dynamics (MD) simulations. Our results suggest that MTH1880 undergoes a two-state N to D transition and it is extremely stable against temperature and denaturants. The reversibility of refolding was confirmed by spectroscopic methods and size exclusion chromatography. We found that the hyper-stability of the thermophilic MTH1880 protein originates from an extensive network of both electrostatic and hydrophobic interactions coordinated by the central ß-sheet. Spectroscopic measurements, in combination with computational simulations, have helped to clarify the thermodynamic and structural basis for hyper-stability of the novel thermophilic protein MTH1880.

Molecular Basis of the Membrane Interaction of the ß2e Subunit of Voltage-Gated Ca(2+) Channels.

The auxiliary ß subunit plays an important role in the regulation of voltage-gated calcium (CaV) channels. Recently, it was revealed that ß2e associates with the plasma membrane through an electrostatic interaction between N-terminal basic residues and anionic phospholipids. However, a molecular-level understanding of ß-subunit membrane recruitment in structural detail has remained elusive. In this study, using a combination of site-directed mutagenesis, liposome-binding assays, and multiscale molecular-dynamics (MD) simulation, we developed a physical model of how the ß2e subunit is recruited electrostatically to the plasma membrane. In a fluorescence resonance energy transfer assay with liposomes, binding of the N-terminal peptide (23 residues) to liposome was significantly increased in the presence of phosphatidylserine (PS) and phosphatidylinositol 4,5-bisphosphate (PIP2). A mutagenesis analysis suggested that two basic residues proximal to Met-1, Lys-2 (K2) and Trp-5 (W5), are more important for membrane binding of the ß2e subunit than distal residues from the N-terminus. Our MD simulations revealed that a stretched binding mode of the N-terminus to PS is required for stable membrane attachment through polar and nonpolar interactions. This mode obtained from MD simulations is consistent with experimental results showing that K2A, W5A, and K2A/W5A mutants failed to be targeted to the plasma membrane. We also investigated the effects of a mutated ß2e subunit on inactivation kinetics and regulation of CaV channels by PIP2. In experiments with voltage-sensing phosphatase (VSP), a double mutation in the N-terminus of ß2e (K2A/W5A) increased the PIP2 sensitivity of CaV2.2 and CaV1.3 channels by ∼3-fold compared with wild-type ß2e subunit. Together, our results suggest that membrane targeting of the ß2e subunit is initiated from the nonspecific electrostatic insertion of N-terminal K2 and W5 residues into the membrane. The PS-ß2e interaction observed here provides a molecular insight into general principles for protein binding to the plasma membrane, as well as the regulatory roles of phospholipids in transporters and ion channels.

Analysis of phosphoinositide-binding properties and subcellular localization of GFP-fusion proteins.

Specific protein-phosphoinositide (PI) interactions are known to play a key role in the targeting of proteins to specific cellular membranes. Investigation of these interactions would be greatly facilitated if GFP-fusion proteins expressed in mammalian cells and used for their subcellular localization could also be employed for in vitro lipid binding. In this study, we found that lysates of cells overexpressing GFP-fusion proteins could be used for in vitro protein-PI binding assays. We applied this approach to examine the PI-binding properties of Aplysia Sec7 protein (ApSec7) and its isoform ApSec7(VPKIS), in which a VPKIS sequence is inserted into the PH domain of ApSec7. EGFP-ApSec7 but not EGFP-ApSec7(VPKIS) did specifically bind to PI(3,4,5)P3 in an in vitro lipid-coated bead assay. Overexpression of EGFP-ApSec7 but not EGFP-ApSec7(VPKIS) did induce neurite outgrowth in Aplysia sensory neurons. Structure modeling analysis revealed that the inserted VPKIS caused misfolding around the PI(3,4,5)P3-binding pocket of ApSec7 and disturbed the binding of PI(3,4,5)P3 to the pleckstrin homology (PH) domain. Our data indicate that plasma membrane localization of EGFP-ApSec7 via the interaction between its PH domain and PI(3,4,5)P3 might play a key role in neurite outgrowth in Aplysia.

Genetic analysis and molecular mapping of low amylose gene du12(t) in rice (Oryza sativa L.).

KEY MESSAGE: We obtained interesting results for genetic analysis and molecular mapping of the du12(t) gene. Control of the amylose content in rice is the major strategy for breeding rice with improved quality. In this study, we conducted genetic analysis and molecular mapping to identify the dull gene in the dull rice, Milyang262. A single recessive gene, tentatively designated as du12(t), was identified as the dull gene that leads to the low amylose character of Milyang262. To investigate the inheritance of du12(t), genetic analysis on an F2 population derived from a cross between the gene carrier, Milyang262, and a moderate amylose content variety, Junam, was conducted. A segregation ratio of 3:1 (χ (2) = 1.71, p = 0.19) was observed, suggesting that du12(t) is a single recessive factor that controls the dull character in Milyang262. Allelism tests confirmed that du12(t) is not allelic to other low amylose controlling genes, wx or du1. Recessive class analysis was performed to localize the du12(t) locus. Mapping of du12(t) was conducted on F2 and F3 populations of Baegokchal/Milyang262 cross. Linkage analysis of 120 F2 plants revealed that RM6926 and RM3509 flank du12(t) at a 2.38-Mb region. To refine the du12(t) locus position, 986 F2 and 289 F3 additional normal plants were screened by the flanking markers. Twenty-six recombinant plants were identified and later genotyped with four additional adjacent markers located between RM6926 and RM3509. Finally, du12(t) was mapped to an 840-kb region on the distal region of the long arm of chromosome 6, delimited by SSR markers RM20662 and RM412, and co-segregated by RM3765 and RM176.