Analysis of stress in the active site of myosin accompanied by conformational changes in transient state intermediate complexes using photoaffinity labeling and 19F-NMR spectroscopy

Myosin forms stable ternary complexes with ADP and the phosphate analogues, fluoroaluminate (Al F4-), fluoroberyllate (BeFn) or orthovanadate (Vi); these ternary complexes mimic transient intermediates in the myosin ATPase cycle. Moreover, we previously demonstrated that these complexes may mimic different myosin ATPase reaction intermediates corresponding to separate steps in the cross-bridge cycle [Maruta, S., Henry, G. D., Sykes, B. D. & Ikebe, M. (1993) J. Biol. Chem. 268, 7093-7100]. Park et al. suggested that the changing conformation of ATP during hydrolysis stresses the active site of myosin subfragment-1 (S-1) through protein-nucleotide contacts at the gamma-phosphate and nucleotide base, and the stress-induced strain in the cross-bridge may be the mechanism by which energy in ATP is transferred to the myosin structure [Park, S., Ajtai, K. & Burghardt, T. P. (1997) Biochemistry 36, 3368-3372]. In the present study, the photoactive ADP analogue, 3'-O-(N-methylanthraniloyl)-2-azido-ADP (Mant-2-N3-ADP), and the 19F-labeled ADP analogue, 2-[(trifluoromethylnitrophenyl)aminoethyl]diphosphate, were employed to examine conformational differences in protein-nucleotide contact in the ATP-binding site that may correlate with energy transduction. Mant-2-N3-ADP was trapped within the active site of skeletal and smooth muscle myosin in the presence of AlF4-, BeFn or Vi. For both skeletal and smooth muscle myosins, trapped Mant-2-N3-ADP was covalently linked to the 25-kDa N-terminal fragment of S-1 of both myosin/Mant-2-N3-ADP/AlF4- and BeFn complexes, presumably at Trp130. However, the efficiency of the incorporation was much higher for skeletal than for smooth muscle myosin suggesting that the conformations of the adenine-binding pockets of the two myosins are somewhat different. Although the amount of Mant-2-N3-ADP trapped in the presence of AlF4- and BeFn was the same for both myosins, the efficiency of photolabeling skeletal muscle myosin was approximately two times higher for BeFn complex than for AlF4- complex. The 19F-NMR spectra of the bound 2-[(trifluoromethylnitrophenyl)aminoethyl]diphosphate in the ternary complexes formed in the presence of AlF4-, BeFn or Vi showed small but distinguishable differences. Taken together, these results indicate that there is some variation in the protein-nucleotide contacts at the nucleotide base among the ternary complexes studied, and these differences mimic separate steps occurring transiently during the contractile cycle.     

Optimal dispersion management for a wavelength division multiplexedoptical soliton transmission system

Soliton's frequency shift arising from the pulse collision between wavelength division multiplexed (WDM) channels in an optical transmission line may be reduced by dispersion management technique. The reduction mechanism is due to a cancellation between the shifts in normal and anomalous dispersion fibers. With a proper management of the dispersion, the effect of lumped amplifiers on the frequency shift may also be canceled out. In this paper, we give a detailed analysis of the frequency shift in dispersion managed systems, and explain the reduction mechanism of the shift. We then propose optimal dispersion management schemes by means of minimizing the frequency shift. We also present a statistical analysis of the collision induced timing jitter for a two-channel WDM system     

Study on combustion and ignition characteristics of natural gas components in a micro flow reactor with a controlled temperature profile

Combustion and ignition characteristics of natural gas components such as methane, ethane, propane and n-butane were investigated experimentally and computationally using a micro flow reactor with a controlled temperature profile. Special attention was paid to weak flames which were observed in a low flow velocity region. The observed weak flame responses for the above fuels were successfully simulated by one-dimensional computations with a detailed kinetic model for natural gas. Since the position of the weak flame indicates the ignition characteristics as well as the reactivity of each fuel, the experimental and computational results were compared with research octane number (RON) which is a general index for ignition characteristics of ordinary fuels. At 1 atm, ethane showed the highest reactivity among these fuels, although RON of ethane (115) is between those of methane (120) and propane (112). Since the pressure conditions are different between the present experiment and the general RON test, weak flame responses to the pressure were investigated computationally for these fuels. The order of the fuel reactivity by the reactor agreed with that by RON test when the pressure was higher than 4 atm. Reaction path analysis was carried out to clarify the reasons of the highest reactivity of ethane at 1 atm among the employed fuels in this study. The analysis revealed that C₂H₅ + O₂ ⇔ C₂H₄ + HO₂ is a key reaction and promotes ethane oxidation at 1 atm. The effect of the pressure on the fuel oxidation process in the present reactor was also clarified by the analysis. In addition, weak flame responses to various mixing ratios of methane/n-butane blends were investigated experimentally and computationally. The results indicated a significant effect of n-butane addition in the blends on combustion and ignition characteristics of the blended fuels.     

Stabilized three-stage oxidation of DME/air mixture in a micro flow reactor with a controlled temperature profile

Ignition and combustion characteristics of a stoichiometric dimethyl ether (DME)/air mixture in a micro flow reactor with a controlled temperature profile which was smoothly ramped from room temperature to ignition temperature were investigated. Special attention was paid to the multi-stage oxidation in low temperature condition.Normal stable flames in a mixture flow in the high velocity region, and non-stationary pulsating flames and/or repetitive extinction and ignition (FREI) in the medium velocity region were experimentally confirmed as expected from our previous study on a methane/air mixture. In addition, stable double weak flames were observed in the low velocity region for the present DME/air mixture case. It is the first observation of stable double flames by the present methodology. Gas sampling was conducted to obtain major species distributions in the flow reactor. The results indicated that existence of low-temperature oxidation was conjectured by the production of CH₂O occured in the upstream side of the experimental first luminous flame, while no chemiluminescence from it was seen.One-dimensional computation with detailed chemistry and transport was conducted. At low mixture velocities, three-stage oxidation was confirmed from profiles of the heat release rate and major chemical species, which was broadly in agreement with the experimental results.Since the present micro flow reactor with a controlled temperature profile successfully presented the multi-stage oxidations as spatially separated flames, it is shown that this flow reactor can be utilized as a methodology to separate sets of reactions, even for other practical fuels, at different temperature.     

Hyper expression of kojibiose phosphorylase gene and trehalose phosphorylase gene from Thermoanaerobacter brockii ATCC35047 in Bacillus subtilis and selaginose synthesis utilizing two phosphorylases

The kojibiose phosphorylase (KP) gene and trehalose phosphorylase (TP) gene from Thermoanaerobacter brockii ATCC35047 were intracellularly hyper-expressed under the control of the Bacillus amyloliquefaciens alpha-amylase promoter in Bacillus subtilis. The production yields were estimated to be 2.1 g of KP and 4.9 g of TP per liter of medium. Selaginose, non-reducing trisaccharide, was synthesized from trehalose utilizing the recombinant KP and TP from B. subtilis. Selaginose was not hydrolyzed by salivary amylase, artificial gastric juice, pancreatic amylase, or small intestinal enzymes.     

Dynamic RWA with FWM-induced crosstalk awareness and modulation format conversion in wavelength-routed optical networks

The constraint of signal transmission quality and network resources utilization in transparent optical networks are herein addressed through a representative cost function based upon an impairment-constraint-base routing (ICBR) approach, taking into account link capacity utilization and the nonlinear physical effect, in the primary instance due to Four-Wave Mixing (FWM)-induced crosstalk. The FWM effect is considered one of the most severe physical impairments for the future photonic networks since the accumulation of FWM crosstalk may cause a fatal degradation in the teletraffic network performance. This paper presents an added functionality to the network model design based on all-optical modulation format conversion from conventional on-off keying (OOK) to quadrature phase-shift keying (QPSK) sending more data within the same bandwidth while minimizing the blocking probability. The joint effect of physical impairment (i.e., FWM-induced crosstalk) awareness and two different modulation formats is considered through numerical simulations. The proposed scheme is expected to realize an all-optical transparent interconnection between networks that employ diverse modulation formats. Results show that the proposed FWM-aware Routing and Wavelength Assignment (RWA) algorithm leads to a more realistic system performance while giving a significant improvement on the network performance.     

Biodegradable Porous Microneedles for an Electric Skin Patch

An array of porous microneedles (PMNs) made of biodegradable poly(lactic-co-glycolic acid) (PLGA) is fabricated by a combination of molding and freeze-drying methods. The optimized mixture of PLGA and 1,4-dioxane is poured into a mold of a microneedle array, followed by the freezing and sublimation of the frozen particles of 1,4-dioxane, a procedure that left an interconnecting porous structure in the PLGA with a porosity around 50%. The mechanical strength of the PMN made of PLGA (PLGA-PMN) is reinforced by modification with carboxymethylcellulose (CMC), resulting in sufficient strength enough for insertion into an excised porcine skin. The transdermal resistance is significantly decreased by the CMC-modified PLGA-PMN, which would improve the efficiency and safety of DC current-based transdermal techniques, including the electrical monitoring of the skin condition and iontophoresis for drug delivery and medical diagnosis.     

Oligonucleotides Containing Phenoxazine Artificial Nucleobases: Triplex-Forming Abilities and Fluorescence Properties

1,3-Diaza-2-oxophenoxazine (“phenoxazine”), a tricyclic cytosine analogue, can strongly bind to guanine moieties and improve π–π stacking effects with adjacent bases in a duplex. Phenoxazine has been widely used for improving duplex-forming abilities. In this study, we have investigated whether phenoxazine and its analogue, 1,3,9-triaza-2-oxophenoxazine (9-TAP), could improve triplex-forming abilities. A triplex-forming oligonucleotide (TFO) incorporating a phenoxazine component was found to show considerably decreased binding affinity with homopurine/homopyrimidine double-stranded DNA, so the phenoxazine system was considered not to function as either a protonated cytosine or thymine analogue. Alternatively, a 9-TAP-containing artificial nucleobase developed by us earlier as a new phenoxazine analogue functioned as a thymine analogue with respect to AT base pairs in a parallel triplex DNA motif. The fluorescence of the 9-TAP moiety was maintained even in triplex (9-TAP:AT) formation, so 9-TAP might be useful as an imaging tool for various oligonucleotide nanotechnologies requiring triplex formation.     

Electrostatic probe measurement in an industrial furnace for high-temperature air conditions

Electrostatic probe measurements are reported that identify flame location, displacement speeds of reaction region, and other flame properties within an industrial furnace that is operated with high-temperature preheated air. The electrostatic probe has advantages over other methods when a furnace is operated with high-temperature air. The probe consisted of a fine detection wire and a supporting tube that played a role of the reference electrode. The reaction regions were found to be widely dispersed and weakened as they moved downstream. However, the ion-current signals still included many sharp peaks, perhaps associated with the thin reaction thickness, contrary to the flame structure expected from the high-temperature air combustion. It was also possible to estimate the displacement speeds of reaction region by using the cross-correlation method between two ion current records detected by parallel detection components. The results demonstrate that the electrostatic probe is useful to detect the structure and state of the reaction mode in industrial furnaces even in the presence of high-temperature air combustion.     

Synchronous Tandem Algorithm for 32-kbit/s ADPCM

The synchronous tandem property of nonaccumulation of distortion in tandem-connected ADPCM coders with a 64 kbit/s PCM interface is discussed here. The synchronous tandem algorithm used to provide this property in the steady-state mode is described with a case-by-case analysis, so as to show how the synchronous tandem property is realized in an ADPCM coder. A 32 kbit/s ADPCM coder utilizing this algorithm has been standardized by the CCITT (International Telegraph and Telephone Consultative Committee). The synchronous tandem property of the 32 kbit/s ADPCM coder is of great interest in network applications, because the ADPCM coder appears likely to be introduced into digital networks built partially with existing 64 kbit/s PCM circuits.