MgATP-induced conformational changes in a single myosin molecule observed by atomic force microscopy: periodicity of substructures in myosin rods

This paper discusses the conformational changes in a single myosin molecule directly observed using atomic force microscopy (AFM). The myosin molecules were pretreated in rigor solutions without MgATP or in relaxed solutions with various concentrations of MgATP. The images of these molecules were obtained using a tapping mode AFM. The results indicate that the orientation of the myosin's heads and tail strongly depend on the MgATP concentration. Without using MgATP, almost all of the myosin molecules are in the extended form; however, when MgATP is used, the molecules bend according to the level of MgATP concentration. The mean-square end-to-end distance of the myosin molecules is significantly shorter with p[MgATP] = 4 than with p[MgATP] = 6. The rod region did not show the same level of intensity along their length in the extended form. The rods exhibited clusters of discontinuity, which were identified as substructures. The size of these substructures change at intervals that are multiples of 14.3-14.5 nm, which reflects the periodicity of the alpha-helical coiled coils. The substructure clusters also correspond to the myosin crossbridge spacing in muscles (14.3 or 43 nm). These results suggest that the myosin's head bends in conjunction with the bending or tilting in the helical substructures. Conformational changes of the myosin molecule induced by MgATP seem to mimic the molecular motions in a muscle's force generation process.     

Strength of bond to bone and cytotoxicity of sintered bodies of hydroxyapatite/zirconia composite particles

Regarding sintered bodies of hydroxyapatite (HAP)/zirconia (PSZ) composite particles prepared by covering the surface of HAP particles with PSZ particles, their strength of the bond to bone after implanting them into an organism and their cytotoxicity were evaluated. Cytotoxicity tests were conducted by the colony formation method. Cytotoxicity was not observed in the sintered bodies of HAP/PSZ composite particles, or in sintered monolithic HAP body and sintered monolithic PSZ body. The strength of bond between a sintered body and a bone was evaluated by measuring the shear strength at the interface between them after a fixed period following implantation of a sintered body into a rabbit femur. In all cases of the sintered monolithic HAP body, and the sintered bodies of HAP/PSZ composite particles and monolithic PSZ body, when the implantation period increased, the shear strength of the sintered body/bone interface tended to increase. In particular, this tendency was very high in the case of sintered bodies of HAP/PSZ with weight ratios of 1/1.0 and 1/1.5, the shear strength for each reaching 20 MPa 12 months after implantation.     

Computer-aided design of a factor Xa inhibitor by using MCSS functionality maps and a CAVEAT linker search

We have investigated a new approach to efficiently find a novel inhibitor against a serine protease (i.e. an activated coagulation factor X, FXa) by using de novo design programs and the X-ray crystal structure of the target enzyme. FXa is a coagulant enzyme that generates thrombin (a serine protease) and participates in both intrinsic and extrinsic coagulation pathways. We adopted multiple copy simultaneous search (MCSS) and CAVEAT linker search techniques, which disclosed a novel FXa inhibitor (T01312) consisting of two binding moieties (the benzamidinyl and adamantyl groups) and a linker unit (the carboxybenzylamine group). The inhibitory activity of T01312 against FXa was determined to be a small K(i)-value of 48nM, which is two orders of magnitude smaller than that against thrombin. An X-ray crystal analysis of T01312 complexed with trypsin (an analogue of FXa) and docking studies of T01312 with trypsin and FXa showed that: (i) the benzamidinyl group is a predominant binding moiety in the anionic pocket (S1 site) with an asparatic acid residue; (ii) a hydrophobic pocket (S4 site) is the binding site of the adamantyl group; (iii) the carboxylate group of the linker contributes to the selectivity for FXa against thrombin. Thus, the combination of the knowledge of the X-ray crystal structure of the target molecule with MCSS and CAVEAT linker search techniques proved to be an effective hit-finding method that does not require the screening of huge compound libraries.     

An improved procedure of electron microscopic in situ hybridization for detecting adenovirus DNA

Electron microscopic in situ hybridization (EM-ISH) is a useful method in determining the localization of a specific nucleic acid at the ultrastructural level. Since the EM-ISH protocol includes many steps, no standard protocol for EM-ISH is available yet. In this study, we optimized quantitatively the critical conditions with respect to embedding resin, nucleic acid labeling and hybridization reaction time, by using adenovirus-infected cells as the indicator cells. The optimal detection of an adenovirus-specific nucleic acid was obtained by overnight hybridization reaction on sections embedded in Lowicryl K4M resin. Random-primed-labeled probes improved the reactivity. At least 60% of virus particles in paracrystalline arrays was found to contain viral DNA. These arrays in adenovirus-infected cells are useful in evaluating quantitatively the efficiency of protocols of EM-ISH.     

Fine silicon carbide fibers synthesized from polycarbosilane-polyvinylsilane polymer blend using electron beam curing

A fine SiC fiber is synthesized from a polymer blend of polycarbosilane (PCS) and polyvinylsilane (PVS) with electron beam curing under vacuum. The obtained SiC fiber from the PCS-PVS blend polymer has smaller average diameter of 8.5 m than that of 11.8 m from PCS, and shows higher average tensile strength of 3.2 GPa than that of 2.8 GPa from PCS after heat treatment at 1673 K in Ar gas atmosphere. However, the SiC fiber from the polymer blend decreases in tensile strength after heat treatment above 1773 K due to -SiC crystal growth near the fiber surface, because of a small amount of oxygen incorporated in the fiber.     

Characterization of fluorine-doped silicon dioxide films by Raman spectroscopy and Electron-spin resonance

We have measured Raman and Electron-spin resonance (ESR) spectra of fluorine-doped SiO₂ films deposited by two different methods. In high-density plasma (HDP) films, the Raman band at about 490 cm^− 1 becomes drastically stronger as the F/Si ratio increases, whereas the Raman band from threefold ring defect is independent of the F/Si ratio. The unusual increase of the intensity of the 490 cm^− 1 band in HDP films has been interpreted in terms of the existence of Si-Si clusters. From a comparison between Raman spectra of HDP film and plasma chemical vapor deposition using tetraethoxysilane (p-TEOS) film with the same F/Si ratios it has been found that HDP film has more Si-Si bonds and threefold ring defects than p-TEOS film. Furthermore, the polarized Raman spectra in the 810 cm^− 1 bands indicate that inhomogeneous SiO₂ clusters of various sizes should exist in the network structure of HDP film. The result of the ESR measurement shows that HDP films have fewer dangling bonds than p-TEOS films. It is considered that many Si-Si clusters, threefold ring defects, and inhomogeneous SiO₂ cluster sizes, and the few dangling bonds in HDP films give rise to the film properties of low stress, good adhesion with Si substrate, and low water permeation.     

Thermal Degradation Under Pulse Operation in Low-Temperature p-Channel Poly-Si Thin-Film Transistors

We analyzed the heat generation of a low-temperature polycrystalline thin-film transistor in pulse operation and proposed a technique for accurately measuring its thermal temperature in high-frequency operation. From this measurement, we were able to calculate the time constants for heating and radiation for the first time. At a low frequency, the temperature difference between when the pulse was on and off was remarkable. As the frequency was increased, the maximum and minimum temperatures approached each other and became equal at a frequency of approximately 1 kHz. We also measured the degradation in pulse operation and discussed the relationship between the thermal temperature and the degradation in the pulse operation     

100-Gb/s DQPSK Transmission Experiment Without OTDM for 100G Ethernet Transport

In order to realize a future 100-Gb Ethernet (100 GbE) transport, 100-Gb/s transmission without 100-GHz-class electronics and optical time-division-multiplexing technique was demonstrated. By using a differential quadrature phase-shift-keying (DQPSK) modulation format and commercially available electronics, 2- and 50-km transmissions of 100-Gb/s signal were successfully achieved over a standard single mode fiber. The receiver sensitivity, chromatic dispersion, and differential group delay tolerances of 100-Gb/s DQPSK signal were also evaluated. Through these evaluations, the possibility of DQPSK modulation for future 100-GbE transport is verified     

Photo‐reductive fading of disperse azo dyes on nylon substrate: thermochemical analyses of reactions of photo‐induced radicals using the semiempirical molecular orbital method†

When the excited triplet states of disperse azo dyes with nitro groups abstract hydrogen to generate hydrazinyl (from azo groups) and nitrosyl hydroxide radicals (from nitro groups), both the radicals as H‐acceptors carry out azo scission, conversion to nitrogen dioxide via disproportionation reactions and self‐decomposition via rearrangement. A kinetic equation was formulated by the sum of these reactions, which describes the initial rates (K_PA) of reductive fading. The K_PA values were controlled by the rate constants of the reactions of hydrazinyl and nitrosyl hydroxide radicals as H‐acceptors, which were estimated by thermochemical analyses of the reactants, intermediates and end products using the semiempirical molecular orbital PM5 method, and by the concentrations of the reactants: H‐acceptors and H‐donors. The K_PA values observed for 12 dyes were explained semi‐quantitatively by multiple routes of reactions depending upon to what extent each radical reaction was thermochemically favoured.