Isolation,Purification and Characterization of a Surfactants-, Laundry Detergents- and Organic Solvents-Resistant Alkaline Protease from <Emphasis Type="Italic">Bacillus</Emphasis> sp. HR-08

Bacillus sp. HR-08 screened from soil samples of Iran, is capable of producing proteolytic enzymes. 16S rDNA analysis showed that this strain is closely related to Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, Bacillus mojavensis, and Bacillus atrophaeus. The zymogram analysis of the crude extract revealed the presence of five extracellular proteases. One of the proteases was purified in three steps procedure involving ammonium sulfate precipitation, DEAE-Sepharose ionic exchange and Sephacryl S-200 gel filtration chromatography. The molecular mass of the enzyme on SDS-PAGE was estimated to be 29 kDa. The protease exhibited maximum activity at pH 10.0 and 60 °C and was inhibited by PMSF but it was not affected by cysteine inhibitors, suggesting that the enzyme is a serine alkaline protease. Irreversible thermoinactivation of enzyme was examined at 50, 60, and 70 °C in the presence of 10 mM CaCl₂. Results showed that the protease activity retains more than 80% and 50% of its initial activity after incubation for 30 min at 60 and 70 °C, respectively. This enzyme had good stability in the presence of H₂O₂, nonionic surfactant, and local detergents and its activity was enhanced in the presence of 20% of dimethyl sulfoxide (DMSO), dimethyl formamide (DMF) and isopropanol. The enzyme retained more than 90% of its initial activity after pre-incubation 1 h at room temperature in the presence of 20% of these solvents. Also, activation can be seen for the enzyme at high concentration (50%, v/v) of DMF and DMSO.     

Engineering of a <Emphasis Type="Italic">Bacillus</Emphasis> α-Amylase with Improved Thermostability and Calcium Independency

Successful industrial use of amylases requires that they are sufficiently stable and active at application conditions, e.g., at high temperature in starch-liquefaction process. In the present study, site-directed mutagenesis was used to enhance the thermal stability and calcium independency of a mesophilic α-amylase from Bacillus megaterium WHO. Mutations (A53S and H58I) were designed at the calcium-binding site based on the sequence alignment. Kinetic and thermostability parameters of the mutants were analyzed and compared with that of the wild type. In the presence of calcium, the affinity of the enzymes (wild type and mutants) toward starch was increased. In comparison to the wild type, calcium ion had more effect on the catalytic efficiency, k _cat/K _m, and half-life (at 60 °C) of A53S mutant. In A53S, the dependence of half-life on calcium concentration showed that the enhanced calcium binding is likely to be responsible for the increased stability. In contrast, calcium-independent mutant (H58I) possessed high thermostability. In addition, thermodynamic parameters of amylolytic reaction exhibited an increase in the activation energy and the entropy of the system. Kinetics of irreversible thermal inactivation suggests that the activation energy increased by 1.4-fold in the most stable variant.     

Multi-domain Fourier-continuation/WENO hybrid solver for conservation laws

We introduce a multi-domain Fourier-continuation/WENO hybrid method (FC–WENO) that enables high-order and non-oscillatory solution of systems of nonlinear conservation laws, and which enjoys essentially dispersionless, spectral character away from discontinuities, as well as mild CFL constraints (comparable to those of finite difference methods). The hybrid scheme employs the expensive, shock-capturing WENO method in small regions containing discontinuities and the efficient FC method in the rest of the computational domain, yielding a highly effective overall scheme for applications with a mix of discontinuities and complex smooth structures. The smooth and discontinuous solution regions are distinguished using the multi-resolution procedure of Harten [J. Comput. Phys. 115 (1994) 319–338]. We consider WENO schemes of formal orders five and nine and a FC method of order five. The accuracy, stability and efficiency of the new hybrid method for conservation laws is investigated for problems with both smooth and non-smooth solutions. In the latter case, we solve the Euler equations for gas dynamics for the standard test case of a Mach three shock wave interacting with an entropy wave, as well as a shock wave (with Mach 1.25, three or six) interacting with a very small entropy wave and evaluate the efficiency of the hybrid FC–WENO method as compared to a purely WENO-based approach as well as alternative hybrid based techniques. We demonstrate considerable computational advantages of the new FC-based method, suggesting a potential of an order of magnitude acceleration over alternatives when extended to fully three-dimensional problems.     

Limited Proteolysis of Luciferase as a Reporter in Nanosystem Biology: A Comparative Study

Firefly luciferase is a 62 kDa protein that produces a flash of light on the oxidation of luciferin in the presence of ATP, Oxygen and Mg²⁺. Luciferase has a broad range of applications for analytical purposes and in vivo imaging for nanosystem biology studies. However, the enzyme is highly susceptible to proteolytic degradation that reduces its half-life. Rate of proteolytic digestion between two members of luciferase family ( Photinus pyralis and Lampyris turkestanicus ) is compared. Proteolytic sensitivity of L. turkestanicus luciferase was found to be more than P. pyralis luciferase, due to higher rate of hydrolysis under identical conditions. Both luciferases showed more sensitivity to chymotrypsin than trypsin with different digestion pattern. Digestion of P. pyralis by trypsin produced some fragments which were found to be more resistant to further degradation, whereas in L. turkestanicus initial fragments subdigested by trypsin, like chymotrypsin effect on both luciferases. Furthermore, both luciferases become increasingly labile to proteolysis as the temperature increases. The rate of inactivation and the rate of degradation between luciferases were different in a specific time of incubation. Appearance of similar bands for both luciferases confirmed exposure of specific regions, in spite of structural differences.     

Evaluation of antiviral activities of curcumin derivatives against HSV-1 in Vero cell line

Antiviral drug resistance is one of the most common problems in medicine, and, therefore, finding new antiviral agents, especially from natural resources, seems to be necessary. This study was designed to assay the antiviral activity of curcumin and its new derivatives like gallium-curcumin and Cu-curcumin on replication of HSV-1 in cell culture. The research was performed as an in vitro study in which the antiviral activity of different concentrations of three substances including curcumin, Gallium-curcumin and Cu-curcumin were tested on HSV-1. The cytotoxicity of the tested compounds was also evaluated on the Vero cell line. The CC50 values for curcumin, gallium-curcumin and Cu-curcumin were 484.2 microg/mL, 255.8 microg/mL and 326.6 microg/mL, respectively, and the respective IC50 values 33.0 microg/mL, 13.9 microg/mL and 23.1 microg/mL. The calculated SI values were 14.6, 18.4 and 14.1, respectively. The results showed that curcumin and its new derivatives have remarkable antiviral effects on HSV-1 in cell culture.     

Synthesis of new 2-substituted pyrazolo[5,1-b][1,3]oxazoles via Sonogashira coupling reactions in water

New 2-substituted pyrazolo[5,1-b][1,3]oxazoles are synthesized through the Sonogashira coupling reaction of 2-iodomethyl-6-methylpyrazolo[5,1-b][1,3]oxazole with terminal alkynes in the presence of a palladium catalyst. This process is carried out in the absence of any copper salt, and provides an eco-friendly and efficient method for the synthesis of the products in water in good-to-high yields. Some of the compounds synthesized were screened for their in vitro anti-oxidant activity using the DPPH (2,2-diphenyl-2-picrylhydrazyl) assays.     

Kinetics and Mechanism Study of the Substitution Reactions of the Chloride Ligand in Dichloro(5,10,15,20)‐tetraphenyl‐21H,23H‐porphinato)tin(IV) by Organic Bases in Dimethylformamide Solvent

Substitution reactions of a Cl⁻ ligand in [SnCl₂(tpp)] (tpp=5,10,15,20‐tetraphenyl‐21H,23H‐porphinato(2−)) by five organic bases i.e., butylamine (BuNH₂), sec‐butylamine (^sBuNH₂), tert‐butylamine (^tBuNH₂), dibutylamine (Bu₂NH), and tributylamine (Bu₃N), as entering nucleophile in dimethylformamide at I=0.1M (NaNO₃) and 30–55° were studied. The second‐order rate constants for the substitution of a Cl⁻ ligand were found to be (36.86±1.14)⋅10⁻³, (32.91±0.79)⋅10⁻³, (22.21±0.58)⋅10⁻³, (19.09±0.66)⋅10⁻³, and (1.36±0.08)⋅10⁻³ M ⁻¹s⁻¹ at 40° for BuNH₂, ^tBuNH₂, ^sBuNH₂, Bu₂NH, and Bu₃N, respectively. In a temperature‐dependence study, the activation parameters ΔH^≠ and ΔS^≠ for the reaction of [SnCl₂(tpp)] with the organic bases were determined as 38.61±4.79 kJ mol⁻¹ and −150.40±15.46 J K⁻¹mol⁻¹ for BuNH₂, 40.95±4.79 kJ mol⁻¹ and −143.75±15.46 J K⁻¹mol⁻¹ for ^tBuNH₂, 30.88±2.43 kJ mol⁻¹ and −179.00±7.82 J K⁻¹mol⁻¹ for ^sBuNH₂, 26.56±2.97 kJ mol⁻¹ and −194.05±9.39 J K⁻¹mol⁻¹ for Bu₂NH, and 39.37±2.25 kJ mol⁻¹ and −174.68±7.07 J K⁻¹ mol⁻¹ for Bu₃N. From the linear rate dependence on the concentration of the bases, the span of k₂ values, and the large negative values of the activation entropy, an associative (A) mechanism is deduced for the ligand substitution.     

Moiré fringes as two-dimensional autocorrelation of transmission function of linear gratings and its application for modulation transfer function measurement

In this work, we show that the transmission moiré pattern is two-dimensional autocorrelation of transmission function (2D-ACTF) of two superimposed linear gratings that make a small angle with each other. In this approach, the properties of moiré fringes vs. the relative displacement of the gratings are formulated. By measuring 2D-ACTF of two similar gratings, we evaluate the modulation transfer function (MTF) of the gratings’ generating system. This work is the generalized form of our previous works. The implied MTF measurement method is easier and faster than our previous methods.     

Nanopolishing by colloidal nanodiamond in elastohydrodynamic lubrication

In this paper, the feasibility of using explosion synthesized diamond nanoparticles with an average particle size (APS) of 3–5 nm with a concentration of 1 % by weight for improving lubrication and friction in elastohydrodynamic lubrication (EHL) was investigated. Owing to the orders of magnitude increase in the viscosity of the lubricant in the EHL contact zone, diamond nanoparticles in the lubricant polish the surfaces at the nanoscale which decreases the composite roughness of contacting surfaces. The reduced composite roughness results in an increased film thickness ratio which yields lower friction. In the numerical analysis, governing equations of lubricant flow in the full elastohydrodynamic lubrication were solved, and the shear stress distribution over the fluid film was calculated. Using an abrasion model and the shear stress distribution profile, the material removal by the nanofluid containing nanoparticles and the resultant surface roughness were determined. The numerical analysis showed that in full EHL regime, the nanolubricant can reduce the composite roughness of moving surfaces. Experimental results from prior studies which exhibited surface polishing by such nanolubricants in boundary, mixed, and full elastohydrodynamic lubrication were used for comparison to the numerical model.     

On dual Bernstein polynomials and stochastic fractional integro-differential equations

In recent years, random functional or stochastic equations have been reported in a large class of problems. In many cases, an exact analytical solution of such equations is not available and, therefore, is of great importance to obtain their numerical approximation. This study presents a numerical technique based on Bernstein operational matrices for a family of stochastic fractional integro-differential equations (SFIDE) by means of the trapezoidal rule. A relevant feature of this method is the conversion of the SFIDE into a linear system of algebraic equations that can be analyzed by numerical methods. An upper error bound, the convergence, and error analysis of the scheme are investigated. Three examples illustrate the accuracy and performance of the technique.