ABSTRACT
Background: Amongst the methods that remove heavy metals from environment, biosorption approaches have received increased attention because of their environmentally friendly and cost-effective feature, as well as their superior performances
Methods: In the present study, we investigated the ability of a surface-engineered Escherichia coli, carrying the cyanobacterial metallothionein on the cell surface, in the removal of Ca [II] from solution under different experimental conditions. The biosorption process was optimized using central composite design. In parallel, the kinetics of metal biosorption was studied, and the rate constants of different kinetic models were calculated
Results: Cadmium biosorption is followed by the second-order kinetics. Freundlich and Langmuir equations were used to analyze sorption data; characteristic parameters were determined for each adsorption isotherm. The biosorption process was optimized using the central composite design. The optimal cadmium sorption capacity [284.69 nmol/mg biomass] was obtained at 40[degree]C [pH 8] and a biomass dosage of 10 mg. The influence of two elutants, EDTA and CaCl[2], was also assessed on metal recovery. Approximately, 68.58% and 56.54% of the adsorbed cadmium were removed by EDTA and CaCl[2] during desorption, respectively. The Fourier transform infrared spectrophotometer [FTIR] analysis indicated that carboxyl, amino, phosphoryl, thiol, and hydroxyl are the main chemical groups involved in the cadmium bioadsorption process
Conclusion: Results from this study implied that chemical adsorption on the heterogeneous surface of E. coli E and optimization of adsorption parameters provides a highly efficient bioadsorbent
ABSTRACT
Background: Improved cyan fluorescent protein [ICFP] is a monochromic, green fluorescent protein [GFP] derivative produced by Aequorea macrodactyla in a process similar to GFP. This protein has strong absorption spectra at wavelengths 426-446 nm. ICFP can be used in cell, organelle or intracellular protein labeling, investigating the protein-protein interactions as well as assessing the promoter activities
Methods: In our previous study, the promoters of two chitinases [ChiS and ChiL] from Bacillus pumilus SG2 were assessed in B. subtilis and their regulatory elements were characterized. In the present study, icfp was cloned downstream of several truncated promoters obtained in the former study, and ICFP expression was evaluated in B. subtilis
Results: Extracellular expression and secretion of ICFP were analyzed under the control of different truncated versions of ChiSL promoters grown on different media. Results from SDS-PAGE and fluorimetric analyses showed that there were different expression rates of CFP; however, the UPChi-ICFP3 construct exhibited a higher level of expression and secretion in the culture medium
Conclusion: Our presented results revealed that inserting this truncated form of Chi promoter upstream of the ICFP, as a reporter gene, in B. subtilis led to an approximately ten fold increase in ICFP expression
Subject(s)
Bacillus subtilis , Chitinases , Bacillus pumilus , DNA, Recombinant , Plasmids , Oligonucleotides , Electrophoresis, Polyacrylamide GelABSTRACT
Objective: Organophosphorus [OPs] compounds are widely used in many pesticides, insecticides and chemical nerve agents. These compounds are hazardous for humans and the environment. Organophosphate hydrolase [OPH] is a homodimeric protein initially isolated from Pseudomonas diminuta MG and Flavobacterium species. This enzyme is able to degrade a broad spectrum of toxic OPs compounds. Using immobilized OPH commonly presents a variety of advantages versus the free form of the enzyme. Advantages include an increase in stability, cost reduction by simple recovery and reutilization of the enzyme, quick and easy separation of the reactant and product in the reaction medium
Methods: Plasmid pET-26b [+] was used to generate the OPH protein under the control of the T7lac promoter. E. coli BL21 [DE3] pLysS was used as the host for expression of the OPH enzyme. Recombinant OPH was secreted into the extracellular medium and the purified enzyme was immobilized on the surface of Bacillus subtilis spores by the adsorption method, for the first time
Results: Approximately 42% to 45% enzymatic activity was determined to be associated with spores. Optimal pH and temperature of the enzyme were not altered by the presence of the spores. Thermo and pH stabilities of the immobilized enzyme was higher than the free form of the enzyme. Conclusion: Bacillus subtilis spores are safe for humans and the environment. Therefore this system can be considered an environmentally friendly biocatalyst for degradation of OPs
Conclusion: Bacillus subtilis spores are safe for humans and the environment. Therefore this system can be considered an environmentally friendly biocatalyst for degradation of OPs.
ABSTRACT
Protein A is a commercially important protein in biotechnological and medicinal applications. The great value of this protein and its applications in genetic and protein engineering and microbial researches as well as the growing use in biochemical industries, biotechnology, medicine and pharmacology, highlight the importance of the present study. In this survey the encoding genes of full-length and truncated forms of protein A were expressed in E. coli under an optimized expression condition. Optimization of the culture conditions resulted in an increase in expression and secretion of both forms of the protein, the pattern of expression and secretion levels for two forms was completely different. A minimum of 10-fold higher expression was observed for the truncated protein in comparison to that of the full-length recombinant form. Hydropathy plot of both forms of proteins showed that the missing domains in the truncated form contain groups of amino acids with high hydrophobicity score. Deletion of the terminal region could led to a higher expression level of the recombinant protein in E. coli. The function of these two proteins was studied using ELISA, which showed a higher activity for the truncated form for binding to IgG, compared to the full-length protein
ABSTRACT
Background: Chitin is an abundant natural polysaccharide found in fungi, algae, and exoskeleton of insects. Several bacterial species are capable of utilizing chitin as their carbon source. These bacteria produce chitinases for degradation of chitin into N-acetyl-D-glucosamine. So far, regulation of the chitinase encoding genes has been studied in different bacterial species. Among Bacillus species, B. pumilus strain SG2 encodes two chitinases, ChiS and ChiL. The promoter region of chiSL genes [PchiS] is mainly regulated by the general carbon catabolite repression [CCR] system in B. subtilis due to the presence of a catabolite responsive element [cre]
Objectives: Use of P[chiS] in constructing an inducible expression system in B. subtilis was investigated
Materials and Methods: In the first step, complete and shortened versions of P[chiS] were inserted upstream of the lacZ on a pBS72/pUC18 shuttle plasmid. The beta-galactosidase activity of B. subtilis carrying one of the relevant plasmids was measured in the presence of different carbon sources
Results: An expression system based on the chitinase promoter of B. pumilus SG2 was established. Modification of PchiS and the culture medium resulted in production of beta-galactosidase in B. subtilis up to 1,800 Miller unit [MU] activity
Conclusions: The chitinase promoter developed in this study, has potential to be used in an expression vector that could be induced by chitin. In addition, compared to the other inducers like IPTG and lactose, chitin is definitely cheaper and more available as an inducer
ABSTRACT
Our preliminary data on the protein expression of SORT1 in ovarian carcinoma tissues showed that sortilin was overexpressed in ovarian carcinoma patients and cell lines, while non-malignant ovaries expressed comparably lower amount of this protein. In spite of diverse ligands and also different putative functions of sortilin [NTR3], the function of overexpressed sortilin in ovarian carcinoma cells is an intriguing subject of inquiry. The aim of this study was, therefore, to investigate the functional role of sortilin in survival of ovarian carcinoma cell line. Expression of sortilin was knocked down using RNAi technology in the ovarian carcinoma cell line, Caov-4. Silencing of SORT1 expression was assessed using real-time qPCR and Western blot analyses. Apoptosis induction was evaluated using flow cytometry by considering annexin-V FITC binding. [3H]-thymidine incorporation assay was also used to evaluate cell proliferation capacity. Real-time qPCR and Western blot analyses showed that expression of sortilin was reduced by nearly 70-80% in the siRNA transfected cells. Knocking down of sortilin expression resulted in increased apoptosis [27.5 +/- 0.48%] in siRNA-treated ovarian carcinoma cell line. Sortilin silencing led to significant inhibition of proliferation [40.1%] in siRNA-transfected Caov-4 cells as compared to mock control-transfected counterpart [p<0.05]. As it was suspected from overexpression of sortilin in ovarian tumor cells, a cell survival role for sortilin can be deduced from these results. In conclusion, the potency of apoptosis induction via silencing of sortilin expression in tumor cells may introduce sortilin as a potential candidate for developing a novel targeted therapy in patients with ovarian carcinoma
ABSTRACT
Chitin, a linear polysaccharide and is one of the most abundant natural polysaccharides is the main component in numerous of organisms. Chitinase enzymes are essential enzymes catalyzing the conversion of chitin to its monomeric or oligomeric components, therefore, chitinase can used as a natural antifungal agent. The purpose of this study is cloning and expression of recombinant chitinse gene in Bacillus subtilis in the treatment offungal infection effects and replacement with carcinogenic and synthetic products. Chitinase gene of Bacillus pumilus was cloned to integrate vector Pdh5 Through designed the appropriate pimers built construct using dual crossing over for transferred into the amylase gene [amyE] of Bacillus subtilis168; and its expression was induced by IPTG and its function was evaluated. Recombinant construct containing chitinase gene was produced. Gene expression by creating halo around the recombinant bacteria colonies in minimal media including chitin as carbon source was observed. Create a halo containing chitin in the minimal media represents the performance of recombinant chitinase enzyme to catalyze the chitin. Considering antifungal property can be effectively treated as a matter of human fungal infection and safe substance to excretion biological pest and pollutants
ABSTRACT
Cell-surface display is the expression of peptides and proteins on the surface of living cells by fusing them to functional components of cells which are exposed to the environment of cells. This strategy can be carried out using different surface proteins of cells as anchoring motifs and different proteins from different sources as a passenger protein. It is a promising strategy for developing novel whole cell factories. Surface engineered cells have many potential uses ranging from medical to environmental applications. This review focuses on different strategy and applications of microbial surface display