ehsan dehnavi

Over expression of Reteplase enzyme has already been studies in the periplasmic space of Escherichia coli (E. coli). However, the role different factors in its expresssin rate remained to be elucidated. Optical cell density (OD), IPTG concentration, and expression time are highly effective in the protein expression rates. Therefore, we aimed to determine the optimum levels of these factors for reteplase expression using response surface methodology (RSM). The pET21b plasmid was used to sub-clone the designed reteplase gene. Then, the gene was transformed into E. coli BL21 strain. Induction of expression was done by IPTG and analyzed by the SDS page. experiments were designed using the RMS, while the effects of different conditions were evaluated using the Real time-PCR. Sequence optimization removed all undesirable sequences of the designed gene. Transformation into E. coli BL21 was confirmed with an 1152 bp band on the agarose gel. A 39 kDa expression band on the SDS gel confirmed the gene expression. Performing 20 RSM-designed experiments, the optimum levels for IPTG concentration and OD were determined as 0.34mM and 5.6, respectively. Moreover, the optimum level of expression time was demonstrated to be 11.91 hours.The accuracy of the regression model for reteplase overexpression was confirmed by an F-value equal to 25.31 and a meager probability value [(Prob > F) < 0.0001]. The real-time-PCR results indicated that the performed calculations were highly accurate. The obtained results indicate that IPTG concentration, OD, and expression time are significantly involved in the augmentation of recombinant reteplase expression. To the best of our knowledge, this is the first study to assess the combined effect of these factors on reteplase expression. Further RSM-based experiments would bring about new insights regarding the best conditions for reteplase expression.

Phytase is an enzyme that has the ability to break down phytic acid into myoinositol and mineral phosphate, and widely uses as an additive in animal foods. The aim of this study was to achieve a high level of bacterial phytase expression in PET26b expression host.

To generate the recombinant phytase enzyme, the target gene was introduced into the expression vector pET-26b-Phytase, and the recombinant vector was transferred to Escherichia coli BL21 as a host of expression after replication in E. coli DH5α. The SDS-PAGE method was used to isolate recombinant phytase and the amount of produced recombinant protein was investigated. by electrophoresis.

The bacterial phytase enzyme was successfully expressed in Escherichia coli and the molecular weight of recombinant phytase produced at about 85 kDa was estimated. The optimum pH for recombinant bacterial phytase was estimated at 5.5.

The results showed that phytase derived from Escherichia coli due to its low production cost and high production of recombinant phytase is a desirable alternative for use in domestic animal feed industry.

Abnormal angiogenesis is associated with various diseases such as solid tumors and metastasis, retinopathies, and rheumatoid arthritis. VEGF-A is the most important mediator of angiogenesis among all growth factors. The bioactivity of VEGF is mediated by two tyrosine kinase receptors VEGFR-1 and VEGFR-2 present on endothelial cells. VEGF signaling through VEGFR-2 is the major angiogenic pathway that leads to stimulation of endothelial cell ingrowths into the tumor. It comprised of an extracellular portion, a cytoplasmic portion, and a short transmembrane domain. The extracellular portion consists of seven Ig-like domains (D1–D7), of which the 1st to 3rd domains function as ligand-binding sites. In the present work, a soluble recombinant extracellular domain 1-3 of VEGFR-2 was expressed in Pichia pastoris to inhibit the VEGF-induced angiogenesis. The 975 bp DNA fragment containing extracellular domain 1-3 kdr, was designed according to the nucleotide sequence at GenBank and protein sequence at Swiss-Prot. The recombinant secretory expression vector (pPinkαHC/KDR1-3) was constructed and transferred into yeast by electroporation. The high expression transformants were identified through complementation of adenine auxotrophy and cultured. KDR1-3 was expressed under the induction of %1 methanol and confirmed by using SDS-PAGE and western blot techniques. After being purified by affinity chromatography using Ni-NTA resins, binding of expressed product to hVEGF165 was proved by two direct ELISA and ELISA receptor binding assays. The data showed that human VEGFR-2 extracellular domain 1-3 with eukaryotic protein structure, that there is no reported paper about, was successfully expressed.

Streptokinase (SK) is an extracellular protein comprising 414 amino acids with considerable clinical importance as a commonly used thrombolytic agent. Due to its wide spread application and clinical importance designing more efficient SK production platforms worth investigatinginvestigation. In this regard, a synthetic SK gene was optimized and cloned in to pET21b plasmid for periplasmic expression. Response surface methodology was used to design a total of 20 experiments for optimization of IPTG concentration, post-induction period, and cell density of induction (OD600). The optimum levels of the selected parameters were successfully determined to be 0.28 mM for IPTG concentration, 9.889 H for post induction period, and 3.40768 for cell density (OD600). These settings result in 4.14 fold increase in SK production rate of optimum expression conditions (7663 IU/mL) in comparison to the primary expression conditions (1853 IU/mL). Achieving higher yields of SK production in shake flask could lead to more cost effective industrial production of this drug which is the ultimate aim of SK production studies.

Demand Response (DR) is counted as one of the main important components in the smart grids. It means the participation of customers in modifying their consumption patterns at the peak hours in order to reduce costs and improve the network reliability. First, in this paper, different types of the incentive based and price based demand response programs (DRPs) and their subsets are investigated. Then, they are modeled and implemented on the Kermanshah province’s load curve on 22 July 2015. Also, their effects on the price reduction and improving the load curve characteristics and network reliability are presented. Finally, a procedure in order to prioritize DRPs based on the electricity company’s different policies is proposed.

Demand Response Programs (DRPs) play an important role in price reduction and reliability improvement in the electrical grids. On the other hand optimal and exact modeling of DRPs can be so effective in the load curve estimation with the lowest error. In this paper the linear and non-linear models of the incentive and price based DRPs have been developed. Modeling is based on the price elasticity matrix (PEM) and customer’s benefit function. Also, the effects of implementation of DRPs on the load curve characteristics such as the load factor, peak to valley, and peak compensate have been investigated. Finally based on different independent system operator’s (ISO’s) policies, DRPs are prioritized using strategy success indexes (SSI).

Dynamic economic emission dispatch (DEED) is a multi-objective optimization problem by which the generators power output are scheduled over the whole dispatch period in order to minimize the fuel cost and emission. This multi-objective optimization problem should meet the load demand constraint and some non-linear constraints such as the valve-point loading effect, prohibited operating zones, and spinning reserve requirements. In this paper an integrated model of the DEED problem and the emergency demand response program (EDRP) has been presented. In the integrated model, the fuel cost and emission are minimized and the optimal incentive is determined simultaneously. In EDRP which is one of the incentive-based demand response programs, incentives are paid to the customers to motivate them reduce their consumption during peak hours or shift it to off-peak hours. The proposed model (DEED-EDRP) is a non-linear complicated optimization problem which may not be solved by the conventional methods. So, four different population-based meta-heuristic algorithms have been used to solve the combined problem. The proposed model has been applied on a ten units test system. Results show that the proposed model is so effective in reducing the total cost and emission and improving the load curve characteristics.

The objective of dynamic economic emission dispatch (DEED) problem is scheduling of the optimal power outputs of the online generating units over a time horizon by minimizing the fuel cost and emission level simultaneously while satisfying the generators and system constraints such as power balance constraint, ramp-rate, and generation limits. In this paper for a more practical and comprehensive study, in addition to the above constraints, the valve-point effect and spinning reserve constraints have been taken into account too. With considering the above conditions, DEED becomes a complex multi-objective optimization problem with non-convex and non-smooth objective function that traditional methods are not able to solve it. So, in this paper random drift particle swarm optimization (RDPSO) has been used to solve the above problem. In addition, a ten unit test system has been studied to demonstrate the effectiveness of the mentioned algorithm and the results are compared with the other algorithms.

Escherichia coli phytase is an acidic histidine phytase with great specific activity. Pichia pastoris is a powerful system for the heterologous expression of active and soluble proteins which can express recombinant proteins in high cell density fermenter without loss of product yield and efficiently secrete heterologous proteins into the media. Recombinant protein expression is influenced by expression conditions such as temperature, concentration of inducer, and pH. By optimization, the yield of expressed proteins can be increase. Response surface methodology (RSM) has been widely used for the optimization and studying of different parameters in biotechnological processes.. In this study, the expression of synthetic appA gene in P. pastoris was greatly improved by adjusting the expression condition.. The appA gene with 410 amino acids was synthesized by P. pastoris codon preference and cloned in expression vector pPinkα-HC, under the control of AOX1 promoter, and it was transformed into P. pastoris GS115 by electroporation. Recombinant phytase was expressed in buffered methanol-complex medium (BMMY) and the expression was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and enzymatic assay. To achieve the highest level of expression, methanol concentration, pH and temperature were optimized via RSM. Finally, the optimum pH and temperature for recombinant phytase activity was determined.. Escherichia coli phytase was expressed in P. pastoris under different cultivation conditions (post-induction temperature, methanol concentration, and post-induction pH). The optimized conditions by RSM using face centered central composite design were 1% (v/v) methanol, pH = 5.8, and 24.5°C. Under the optimized conditions, appA was successfully expressed in P. pastoris and the maximum phytase activity was 237.2 U/mL after 72 hours of expression.. By optimization of recombinant phytase expression in shake flask culture, we concluded that P. pastoris was a suitable host for high-level expression of phytase and it can possess high potential for industrial applications..

Electricity generation by the wind power in the recent years has increased a lot. The main reason is that it doesn’t use fossil fuels and consequently has no environmental pollution and reduces greenhouse gases. But due to the fact that wind farms are not available at everywhere, so generation by this method is not always economical and reachable. Wind speed is variable and can be destructive, so induction generators are used in wind farms because design, construction, and maintenance of these machines are much easier than synchronous machines. But these generators have some disadvantages too for example they use a lot of reactive power and cause voltage instability. Also due to the unpredictable behavior of the wind speed, output voltage of wind farm is unstable too. One solution to this problem is using of Flexible AC Transmission System (FACTS) for the reactive power compensation. In this article the use of STATCOM has been investigated and analyzed. At first a wind farm in the variable wind speed has been simulated by Matlab (Simulink) and the effects of STATCOM and SVC in the transient and steady state conditions have been investigated and compared. Finally due to the simulation results and previous researches, it has been proved that STATCOM is the best compensator in the wind farm.

β- Xylosidase from Selenomonas ruminantium (SXA) is one of the most important enzyme for the hydrolysis of cell wall hemicellulose. SXA has potential utility in industrial processes especially production of bioethanol from bagasse. However، this xylosidase lose activ ity drastically above 50 °C. Each monomer of this homotetramer has four free buried cysteine. It seems that cysteine 286 has no role in protein function. In this study، to investigate effects of free buried cysteine on protein thermal stability، Cys 286 was replaced with the same size amino acid، valine. The mutant and native protein have expressed in Pichia pastoris. Kinetic and thermostability parameters of mutant were compared with the wild type enzyme. While pH optimum، temperature profile and catalytic efficiency of recombinant mutant were be found similar to native enzyme، mutant showed about 65% increase in thermostability respect to the wild type at 55 ˚C. Our results showed that free thiol group of cysteine caused the destabilization. Moreover، hydrophobic side chain of valine could involve in a hydrophobic interaction to stabilize SXA. Elimination of a free cysteine enhanced thermal stability without changing the catalytic efficiency of the enzyme that could be very important for biotechnological applications.

β-D-Xylosidases (EC 3. 2. 1. 37)، one of the most important hemicellulases with high potential industrial application such as bioethanol production، are exo-type glycosidases that catalyze the successive removal of β-xylosyl residues from the non-reducing termini of xylobiose and higher linear β-1،4-xylooligosaccharides and in conjunction with β-xylanases are essential in completely depolymerizing xylan. The aim of this study was secretive expression of bacterial β-xylosidase gene including hexahistidin-tag in Pichia pastoris in order to achieve high level expression and enzyme purification subsequently. In this study، after optimizing the sequence of protein encoding bacterial β-xylosidase based on the expression codon usage of Pichia pastoris and addition of 6xHis-tag sequence through suitable designed primers to gene، transformation of recombinant plasmids was performed through electroporation method into the expression yeast. The results showed that the recombinant β-xylosidase production was 40 U/L with 0/6 U/mg specific activity in flask culture. Cloning and successful expression of bacterial β-xylosidase gene was carried out in order to obtain an active and stable enzyme with high level expression from yeast expression system