Bacillus subtilis转录因子CtsR蛋白中clpC操纵子结合区域的NMR研究

革兰氏阳性菌枯草芽孢杆菌（Bacillus subtilis）响应热刺激时,精氨酸激酶McsB磷酸化转录因子CtsR蛋白中clpC操纵子结合区域的Arg62（R）位点,使得CtsR与clpC操纵子解离,从而启动clpC相关基因的转录过程,以表达细菌应对热刺激所需的蛋白.本文以CtsR蛋白中结合clpC操纵子的区域（KRGGGG）为研究对象,通过对¹H NMR、¹H-¹H COSY、¹H-¹H TOCSY、¹H-¹⁵N HSQC和¹H-¹³C HSQC等谱图的综合分析,对其¹H、¹³C以及¹⁵N的化学位移进行归属,为该片段与clpC操纵子相互作用,以及精氨酸磷酸化调控机制的研究提供基础.     

Microscopical investigations of nisin-loaded nanoliposomes prepared by Mozafari method and their bacterial targeting

Nanoencapsulation may improve activity of protein or polypeptide antimicrobials against a variety of microorganisms. In this study, nanoliposomes prepared from different lipids (Phospholipon 90H, Phospholipon 100H, dipalmitoylphosphatidylcholine (DPPC), stearylamine (SA), dicetyl phosphate (DCP) and cholesterol) by a new, non-toxic and scalable method, were tested for their capacity to encapsulate nisin Z and target bacteria (Bacillus subtilis and Pseudomonas aeruginosa). Factors affecting the entrapment efficiency (charge and cholesterol concentration in the vesicles) and stability of nanoliposomes were assessed. The nanoliposomes and their bacterial targeting were visualised, using different microscopes under air and liquid environments. Nisin was entrapped in different nanoliposomes with encapsulation efficiencies (EE) ranging from 12% to 54%. Anionic vesicles possessed the highest EE for nisin while increase in cholesterol content in lipid membranes up to 20% molar ratio resulted in a reduction in EE. Stability of nanoliposome-encapsulated nisin was demonstrated for at least 14 months at 4 °C (DPPC:DCP:CHOL vesicles) and for 12 months at 25 °C (DPPC:SA:CHOL vesicles).     

Comparison between the use of polyether ether ketone and stainless steel columns for ultrasonic-assisted extraction under various ultrasonic conditions

The ultrasound-assisted extraction (UAE) was conducted using the stainless steel (SS) and polyether ether ketone (PEEK) columns and analyzed with high-performance liquid chromatography (HPLC) to understand the mechanism of ultrasound-assisted chromatography (UAC). Empty SS and PEEK columns were used to extract dyes from a fabric under identical conditions with several parameters including the initial ultrasonic bath temperatures (30 °C and 40 °C), ultrasound power intensities (0, 20, 40, 60, 80, and 100 %), ultrasound operation modes (normal and sweep), and ultrasound frequencies (25 kHz, 40 kHz, and 132 kHz) to compare their extraction capabilities. After 30 min of extraction, the amount of extract was determined by HPLC. The PEEK material was significantly affected by ultrasonic radiation compared to the SS material, especially at a higher temperature (40 °C), power intensity (100 %), and frequency (132 kHz) with sweep mode. At a maximum power density of 45 W/L, the extraction effectiveness ratio of PEEK to SS was in the range of 1.8 - 3.9 depending on the specific frequency, initial temperature, and with or without temperature control. The most optimal ultrasound frequencies, in terms of enhancing extraction effectiveness, are in the order of 132 kHz, 40 kHz, and 25 kHz. Unlike the SS material, the PEEK material was more affected by temperature and acoustic effects under identical conditions, especially at 132 kHz ultrasound frequency. In contrast, at lower frequencies of 40 kHz and 25 kHz, no significant differences in the acoustic effects were observed between the PEEK and SS materials. The findings of this study contribute to elucidating the roles of column materials in UAE and UAC.     

Dynamic supercoiling bifurcations of growing elastic filaments

Certain bacteria form filamentous colonies when the cells fail to separate after dividing. In Bacillus subtilis, Bacillus thermus, and Cyanobacteria, the filaments can wrap into complex supercoiled structures as the cells grow. The structures may be solenoids or plectonemes, with or without branches in the latter case. Any microscopic theory of these morphological instabilities must address the nature of pattern selection in the presence of growth, for growth renders the problem nonautonomous and the bifurcations dynamic. To gain insight into these phenomena, we formulate a general theory for growing elastic filaments with bending and twisting resistance in a viscous medium, and study an illustrative model problem: a growing filament with preferred twist, closed into a loop. Growth depletes the twist, inducing a twist strain. The closure of the loop prevents the filament from unwinding back to the preferred twist; instead, twist relaxation is accomplished by the formation of supercoils. Growth also produces viscous stresses on the filament which even in the absence of twist produce buckling instabilities. Our linear stability analysis and numerical studies reveal two dynamic regimes. For small intrinsic twist the instability is akin to Euler buckling, leading to solenoidal structures, while for large twist it is like the classic writhing of a twisted filament, producing plectonemic windings. This model may apply to situations in which supercoils form only, or more readily, when axial rotation of filaments is blocked. Applications to specific biological systems are proposed.     

Influence of ultrasound power and frequency upon corrosion kinetics of zinc in saline media

This paper is devoted to zinc corrosion and oxidation mechanism in an ultrasonically stirred aerated sodium sulfate electrolyte. It follows a previous study devoted to the influence of 20 kHz ultrasound upon zinc corrosion in NaOH electrolytes [Ultrason. Sonochemis. 8 (2001) 291]. In the present work, various ultrasound regimes were applied by changing the transmitted power and the wave frequency (20 and 40 kHz). Unlike NaOH electrolyte which turns the zinc electrode into a passive state, Na₂SO₄ saline media induces soft corrosion conditions. This allows a study of the combined effects of ultrasonically modified hydrodynamic and mechanical damage (cavitation) upon the zinc corrosion process. A series of initial experiments were carried out so as to determine the transmitted power and to characterize mass transfer distribution in the electrochemical cell. Zinc corrosion and oxidation process were subsequently studied with respect to the vibrating parameters. When exposed to a 20 kHz ultrasonic field, and provided that the electrode is situated at a maximum mass transfer point, the corrosion rate reaches values six to eight times greater than in silent conditions. The zinc oxidation reaction, in the absence of competitive reduction reactions, is also activated by ultrasound (20 and 40 kHz) but probably through a different process of surface activation.     

The effect of sonication on microbial disinfection using hypochlorite

Ultrasound alone is capable of killing bacteria when sufficient power is applied but ultrasound at low powers can also be used to improve the effectiveness biocides. In this paper, we explore the effect of the timing of the ultrasonic treatment at 20 and 850 kHz on the biocidal efficiency of sodium hypochlorite solution towards Escherchia coli suspensions. A remarkable frequency effect has been noted. At the lower frequency of 20 kHz the improvement in biocidal activity is greatest when the ultrasound is applied at the same time as the hypochlorite. At the higher frequency of 850 kHz the improvement is best when ultrasound is used as a pre-treatment immediately followed by hypochlorite addition under normal (silent) conditions. The kill rate achieved for pre-treatment using 850 kHz and simultaneous treatment using 20 kHz are very similar. However the former involves less acoustic energy and so is considered to be the more efficient.     

Application of ultrasound in preparing pathological sections to reduce processing time

It has been found experimentally that application of sub-mega and low megahertz ultrasound (US) of spatial and temporal averaged intensity I_sata up to 10 W/cm² during the process of preparing pathological sections of the mouse tissue has shortened the processing time from 12 h (without US) to less than half an hour (with US). The experiment has also showed that the processing time reached the shortest for ultrasound f = 200 kHz among the frequencies of 200 kHz, 400 kHz, 600 kHz, 800 kHz and 1 MHz used in this study. It has been proposed that ultrasound inducing non-inertial cavitation enhanced the permeability of cell membrane to liquid. Thus tissue fixation and dehydration were speeded up by application of US.     

Frequency, field, and temperature dependence of the AC penetration depth of a GdBa2Cu3O7−δ film in the mixed state

We report on a systematic mutual induction measurement of the complex AC penetration depth λ of a sputtered high-quality GdBa₂Cu₃O_7−δ film in the mixed state by a very small coil system arranged near the sample surface. The complex penetration depth λ(B, T, ω) for DC inductions B 0.65 T (perpendicular to the film), for temperatures 36 K T 81 K, and for frequencies 1 kHz ω/2π 500 kHz was determined from the measured signal by a novel inversion scheme. The results are consistent with theoretical predictions based upon single vortex pinning. The Labusch parameter , the flux creep relaxation time τ, as well as the effective activation energy U are simulateneously determined.     

In vitro ultrasound-mediated leakage from phospholipid vesicles

Interest in using ultrasound energy in wound management and intracellular drug delivery has been growing rapidly. Development and treatment optimization of such non-diagnostic applications requires a fundamental understanding of interactions between the acoustic wave and phospholipid membranes, be they cell membranes or liposome bilayers. This work investigates the changes in membrane permeation (leakage mimicking drug release) in vitro during exposure to ultrasound applied in two frequency ranges: “conventional” (1 MHz and 1.6 MHz) therapeutic ultrasound range and low (20 kHz) frequency range. Phospholipids vesicles were used as controllable biological membrane models. The membrane properties were modified by changes in vesicle dimensions and incorporation of poly(ethylene glycol) i.e. PEGylated lipids. Egg phosphatidylcholine vesicles with 5 mol% PEG were prepared with sizes ranging from 100 nm to 1 μm. Leakage was quantified in terms of temporal fluorescence intensity changes observed during carefully controlled ultrasound ON/OFF time intervals. Custom-built transducers operating at frequencies of 1.6 MHz (focused) and 1.0 MHz (unfocused) were used, the I_spta of which were 46.9 W/cm² and 3.0 W/cm², respectively. A commercial 20 kHz, point-source, continuous wave transducer with an I_spta of 0.13 W/cm² was also used for comparative purposes. Whereas complete leakage was obtained for all vesicle sizes at 20 kHz, no leakage was observed for vesicles smaller than 100 nm in diameter at 1.6 or 1.0 MHz. However, introducing leakage at the higher frequencies became feasible when larger (greater than 300 nm) vesicles were used, and the extent of leakage correlated well with vesicle sizes between 100 nm and 1 μm. This observation suggests that physico-chemical membrane properties play a crucial role in ultrasound mediated membrane permeation and that low frequency (tens of kilohertz) ultrasound exposure is more effective in introducing permeability change than the “conventional” (1 MHz) therapeutic one. The experimental data also indicate that the leakage level is controlled by the exposure time. The results of this work might be helpful to optimize acoustic field and membrane parameters for gene or drug delivery. The outcome of this work might also be useful in wound management.     

Estimation of hydroxyl free radicals produced by ultrasound in Fricke solution used as a chemical dosimeter

Hydroxyl free radicals produced in Fricke solution exposed to 80 kV X-rays or 23 kHz ultrasound (intensity 3 W cm⁻²) or 20 kHz ultrasound (intensity 18.9 W cm⁻²) or 3.5 MHz clinical ultrasound (intensity 1.47 W cm⁻²), as estimated from the Fricke dosimetric data, exhibited a linear dose-response relationship. The dosimeter was found to be effective in the concentration range 1.0–8.0 mM of FeSO₄ solution. The hydroxyl radicals produced in Fricke solution were inhibited by the OH radical scavengers dimethyl sulfoxide (200 mM), -histidine (10 mM) and sodium benzoate (10 mM) in a manner proportional to the rate constants of their reaction with the OH radicals. The power threshold for OH radical formation, which is presumably the threshold for cavity formation, was estimated for 23 kHz ultrasound by this dosimeter as 1.28 W cm⁻² for a 4 cm³ sample volume.     

Electrical and dielectric characterization of Au/ZnO/n-Si device depending frequency and voltage

Au/Zn O/n-type Si device is obtained using atomic layer deposition(ALD) for Zn O layer, and some main electrical parameters are investigated, such as surface/interface state(Nss), barrier height(Φb), series resistance(Rs), donor concentration(Nd), and dielectric characterization depending on frequency or voltage. These parameters are acquired by use of impedance spectroscopy measurements at frequencies ranging from 10 k Hz to 1 MHz and the direct current(DC) bias voltages in a range from-2 V to +2 V at room temperature are used. The main electrical parameters and dielectric parameters,such as dielectric constant(ε"), dielectric loss(ε"), loss tangent(tan δ), the real and imaginary parts of electric modulus(M and M), and alternating current(AC) electrical conductivity(σ) are affected by changing voltage and frequency. The characterizations show that some main electrical parameters usually decrease with increasing frequency because charge carriers at surface states have not enough time to fallow an external AC signal at high frequencies, and all dielectric parameters strongly depend on the voltage and frequency especially in the depletion and accumulation regions. Consequently, it can be concluded that interfacial polarization and interface charges can easily follow AC signal at low frequencies.     

Effect of ultrasonic irradiation on production of fermented milk with Lactobacillus delbrueckii

Milk fermentation with Lactobacillus delbrueckii under ultrasonic irradiation was carried out in a 450 cm³ bioreactor with a polyethylene film bottom. Ultrasonic irradiation increased the hydrolysis of lactose in milk but decreased the cell viability. However, the viable cell count increased again when the ultrasound was stopped, because ultrasound did not destroy the ability for cell propagation. When the sonication power was 17.2 kW m^-2 and the sonication period was 3 h, 4.9 × 10⁸ cfu cm^-3 of the viable cell count and 55% lactose hydrolysis were attained. In contrast, the viable cell count was 2 × 10⁹ cfu cm^-3 and 35.6% lactose was hydrolysed in control fermentation.     

Synergistic effect of hydrogen peroxide production and sonochemiluminescence under dual frequency ultrasound irradiation

The synergistic effect of H(2)O(2) production and sonochemiluminescence (SCL) was studied under both orthogonal and opposite dual irradiation at the frequencies of 28, 584 and 970 kHz and at various acoustic powers. The largest reduction in H(2)O(2) production was observed under opposite dual irradiation at a 28/28 kHz frequency without considering the acoustic power levels. The largest enhancement was observed under dual irradiation at a frequency of 28/970 kHz. This enhancement might be due to the increased number of bubbles that underwent violent collapse by low frequency ultrasound (28 kHz). These results were also confirmed by observing the SCL. Under dual irradiation at relatively high frequencies (i.e., 584 and 970 kHz), the synergistic effect was high at low acoustic power levels. However, the effect tended to decrease (to the equivalent of the calculation from the result of each single irradiation) with increasing acoustic power. Unlike dual irradiation coupled with a frequency of 28 kHz, the inhibition effect was not observed under dual irradiation at relatively high frequencies. With respect to H(2)O(2) production, the production rate constants of H(2)O(2) followed the order of 584/584>584/970>28/970≈28/584>28/28 kHz, which resulted from the fact that the production efficiency of H(2)O(2) at an irradiation frequency of 584 kHz was considerably higher than that at other frequencies.     

Intensification of leaching process by dual-frequency ultrasound

Ultrasound is gaining importance in metal extraction process. In the previous laboratory scale investigation the authors have established the positive influence of ultrasound on copper recovery from oxide ores of Malanjkhand, Madhya Pradesh, India in an ammonical media. The process parameters in a conventional agitation method were optimized and a maximum recovery of ≈32% in 20 min was obtained without sonication. The recovery was increased to ≈78% by the application of ultrasound over the same period with several advantages like decrease in leaching time and the reagent consumption. In the present study the leaching process is intensified by studying the metal recovery variation at different ultrasonic frequencies (20, 40, 43 and 720 kHz) and intensities (up to 8 W cm⁻²) with sonication time. The results show that sinusoidal ultrasound even at larger intensity has some limitations with single frequency. However, simultaneous application of dual frequency 20 and 40 kHz ultrasound enhanced extraction rates along with increased yield. While conventional single frequency exposure at either one of the two frequencies at the same acoustic power level did not yield similar results, application of two wave sources, as used in the study revealed that it is possible to save energy through lowering of time of operation process.     

Adsorption kinetics of 4-chlorophenol onto granular activated carbon in the presence of high frequency ultrasound

This work describes the results of investigations carried out to examine the adsorption kinetics of 4-chlorophenol (4-CP) from aqueous solution containing tert-butyl alcohol (10%, v/v) onto granular activated carbon (GAC) in the presence of ultrasound of different high frequencies (516, 800 and 1660 kHz) and acoustic powers (15.2, 21.5, 31.1 and 38.3 W). The main objective of this study is to describe the mechanism of ultrasound-assisted adsorption rather than the enhancement of adsorption capacity. Sonochemical degradation of 4-CP was studied in the absence and presence of tert-butyl alcohol. The sonolysis of 4-CP is effectively inhibited by the addition of tert-butyl alcohol (10%, v/v) and very little 4-CP degradation occurs, indicating that little or no pyrolysis of the compound occurs. Without addition of tert-butyl alcohol, after 300 min and at 1660 kHz, the removal of 4-CP in the presence of ultrasound for an acoustic power of 38.3 W was nearly total (99%), but in the conventional method only 60% was eliminated. In this case, the removal of 4-CP by GAC in the ultrasound-assisted technique is due to both adsorption and ultrasonic degradation, but the removal by simple stirring is only due to adsorption, which makes a direct comparison unacceptable. In order to distinguish sonochemical degradation and adsorption of 4-CP onto GAC and to make an exact and practical comparison of the adsorption in the absence and presence of ultrasound, kinetic adsorption experiments were conducted using aqueous solution containing 10% (v/v) tert-butyl alcohol. The obtained results show that both adsorption rate and adsorbed amount were significantly enhanced and improved in the presence of ultrasound for all the studied frequencies and powers. The enhancement of adsorption is favored by increasing ultrasonic power. Adsorption kinetic data were modeled using the liquid-film mass transfer equation and intraparticle diffusion model. The values of the intraparticle diffusion coefficient obtained in the presence of ultrasound are greater than that obtained in the absence of ultrasound. In the initial period of adsorption, where external mass transfer is assumed to predominate, liquid-film mass transfer coefficients significantly increased by the assistance of ultrasound. These results indicate that ultrasound enhances the mass transport in the pores as well as across the boundary layer. This effect increased with increasing ultrasonic power for the three studied frequencies. The average order for the studied ultrasonic waves according to the initial adsorption rate, the intraparticle diffusion coefficient and the liquid-film mass transfer coefficient is 516 kHz>800 kHz>1660 kHz.     

Dosimetry in sonochemistry: the use of aqueous terephthalate ion as a fluorescence monitor

The generation of HO radicals by acoustic cavitation in water was monitored by their reaction with terephthalic acid (TA) anion to produce fluorescent hydroxyterephthalate ions using a cleaning bath (38kHz) and a probe system (20, 40 and 60 kHz) as different sources of ultrasound. When using the ultrasonic bath as a source of energy for sonochemical studies, the shape of the reaction vessel is important. In the case of HO production from water (50 cm³), reaction in a conical flask (100 cm³) produces 2.75 times more radicals than a round-bottomed flask of the same capacity. The fluorescence yield (fluorescence intensity/ultrasound dosage) obtained using the conical flask and ultrasonic bath was similar to that for a probe operating at 40 kHz on the same volume of solution. For a probe system operating at 20, 40 and 60 kHz the greatest sonochemical efficiency was attained at the highest of these frequencies (60 kHz). For the probe system the fluorescence yield is directly proportional to power input and the concentration of TA. The fluorescence yield decreases as the temperature is increased.     

The effect of ultrasound treatment in combination with nisin on the inactivation of Listeria innocua and Escherichia coli

Ultrasound, alone or in combination with natural antimicrobials, is a novel food processing technology of interest to replace traditional food decontamination methods, as it is milder than classical sterilisation (heat treatment) and maintains desirable sensory characteristics. However, ultrasound efficacy can be affected by food structure/composition, as well as the order in which combined treatments are applied. More specifically, treatments which target different cell components could result in enhanced inactivation if applied in the appropriate order. The microbial properties i.e. Gram positive/Gram negative can also impact the treatment efficacy.This work presents a systematic study of the combined effect of ultrasound and nisin on the inactivation of the bacteria Listeria innocua (Gram positive) and Escherichia coli (Gram negative), at a range of cavitation conditions (44, 500, 1000 kHz). The order of treatment application was varied, and the impact of system structure was also investigated by varying the concentration of Xanthan gum used to create the food model systems (0 – 0.5% w/v). Microbial inactivation kinetics were monitored, and advanced microscopy and flow cytometry techniques were utilised to quantify the impact of treatment on a cellular level.Ultrasound was shown to be effective against E. coli at 500 kHz only, with L. innocua demonstrating resistance to all frequencies studied. Enhanced inactivation of E. coli was observed for the combination of nisin and ultrasound at 500 kHz, but only when nisin was applied before ultrasound treatment. The system structure negatively impacted the inactivation efficacy. The combined effect of ultrasound and nisin on E. coli was attributed to short-lived destabilisation of the outer membrane as a result of sonication, allowing nisin to penetrate the cytoplasmic membrane and facilitate cell inactivation.     

Multi-frequency ultrasound:A potential method to improve the effects of surface decontamination and structural characteristics on large yellow croaker (Pseudosciaena crocea) during refrigerated storage

The effects of multi-frequency ultrasound on surface decontamination and structural characteristics of large yellow croaker (Pseudosciaena crocea) during refrigerated storage were evaluated. The results of total viable counts (TVCs) and psychrophilic bacteria counts (PBCs) demonstrated that multi-frequency ultrasound retarded the growth of microorganisms. The bacteriostatic effect was positively correlated with the increase of ultrasound frequencies. However, compared with triple-frequency ultrasound (TUS, 20/28/40 kHz) treatment, dual-frequency ultrasound (DUS, 20/28 kHz) treatment had higher water-holding capacity (WHC) and immobilized water content, better texture characteristics, lower pH and total volatile basic nitrogen (TVB-N). Through the results of myofibrillar fragmentation index (MFI), intrinsic fluorescence intensity (IFI) and atomic force microscope (AFM), multi-frequency ultrasound could effectively stabilize the myofibrillar protein structure of refrigerated large yellow croaker, which could maintain better texture characteristics. The effects of DUS were the most significant. Therefore, multi-frequency ultrasound treatment could inhibit the growth of microorganisms and improve the structural characteristics of large yellow croaker during refrigerated storage.     

On the string interpretation of M(atrix) theory

It has been proposed recently that, in the framework of M(atrix) theory, = 8 supersymmetric U(N) Yang-Mills theory in 1 + 1 dimensions gives rise to type IIA long string configurations. We point out that the quantum moduli space of SYM_{1 + 1} gives rise to two quantum numbers, which fit very well into the M(atrix) theory. The two quantum numbers become familiar if one switches to a IIB picture, where they represent configurations of D-strings and fundamental strings. We argue that, due to the SL(2,Z) symmetry, of the IIB theory, such quantum numbers must represent configurations that are present also in the IIA framework.     

Inactivation of microorganisms by low-frequency high-power ultrasound: 2. A simple model for the inactivation mechanism

A simple theoretical model based on shear forces generated by the collapse of the ultrasound cavities near the surface of a microorganism is proposed. This model requires two parameters which take into account the number of acoustic cavitation bubbles, and the resistance of the cell wall of the microorganism to the shear forces generated by bubble collapse. To validate the model, high-power low frequency (20 kHz) ultrasound was used to inactivate two microorganisms with very different sizes, viz., a bacterium, Enterobacter aerogenes and a yeast, Aureobasidium pullulans. The inactivation ratio was experimentally measured as a function of sonication time for different ultrasound power and for different initial cell numbers. For both E. aerogenes and A. pullulans the Log of the inactivation ratio decreased linearly with sonication time, and the rate of inactivation increased (D-value decreased) with the increase in sonication power. The rate of inactivation was also found, for both microorganisms, to increase with a decrease in the initial cell number. The fits, obtained using the proposed model, are in very good agreement with the experimental data.