Taha Zaghloul - Independent Researcher (original) (raw)

Papers by Taha Zaghloul

Procedia Technology, 2015

A simple assay for determining activities of phosphopentomutase from a hyperthermophilic bacterium Thermotoga maritima

Analytical biochemistry, Jan 15, 2016

Phosphopentomutase (PPM) catalyzes the interconversion of α-D-(deoxy)-ribose 1-phosphate and α-D-... more Phosphopentomutase (PPM) catalyzes the interconversion of α-D-(deoxy)-ribose 1-phosphate and α-D-(deoxy)-ribose 5-phosphate. We developed a coupled or uncoupled enzymatic assay with an enzyme nucleoside phosphorylase for determining PPM activities on D-ribose 5-phosphate at a broad temperature range from 30 to 90 °C. This assay not only is simple and highly sensitive but also does not require any costly special instrument. Via this technology, an open reading frame TM0167 from a thermophilic bacterium Thermotoga maritima putatively encoding PPM was cloned. The recombinant PPM was overexpressed in Escherichia coli Rosetta. This enzyme has the highest activity at 90 °C. MnCl2 (0.1 mM) and 50 μM α-D-glucose 1,6-bisphosphate are cofactors. The kinetic parameters of Km and kcat are 1.2 mM and 185 s(-1) at 90 °C, respectively. The enzyme has a half-life time of up to 156 min at 90 °C. This enzyme is the most active and thermostable PPM reported to date.

Molecular Cloning and Expression in Escherichia coli of Pseudomonas aeruginosa lipase gene

Biotechnology, 2006

Polish Journal of Microbiology Polskie Towarzystwo Mikrobiologow the Polish Society of Microbiologists, Feb 1, 2005

Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x t... more Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x to apparent homogeneity using ammonium sulfate precipitation followed by ion exchange chromatography on Q-and S-Sepharose column. Both of the purified lipases are monomeric protein with molecular weight of 15.5 and 54.97 KDa respectively. The optimal activities of the enzymes were at 45 and 50°C and pHs 10.0 and 9.0. Calcium ions increase thermostability of both purified lipases I and II. The purified lipase I showed no metal ion dependence for its activity since EDTA up to 10 mM has no effect on the enzyme activity. However purified lipase II showed slight inhibition by EDTA at the same concentration. Moreover, a serine protease inhibitor, PMSF showed an inhibitory effect on both purified enzymes. K e y w o r d s: Lipases, Pseudomonas aeruginosa Ps-x, 16S rRNA Abbreviations: SDS-PAGE; sodium dodecyl sulfate polyacrylamide gel electrophoresis, Q-Sepharose; Quaternary amino methyl Sepharose, EDTA; ethylene diamine tetraacetic acid.

The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poult... more The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poultry feather and sheep wool, has made keratinase production by microorganisms highly important to the biotechnological industry. A proteindegrading bacterium (C 4 ) was isolated from compost. Based on morphology and biochemical tests, along with 16S rRNA sequencing, the isolated C 4 was tentatively identified as Bacillus sp. C 4 (2008). The proteolytic activity of the Bacillus sp. C 4 strain was broadly specific; it degraded keratinous and non-keratinous proteins to different degrees. Pea pods as substrate generated the highest protease production, followed by soybean meal and sheep wool. Notwithstanding, using wool keratin as a sole source of carbon and nitrogen yielded the highest level of soluble proteins. Furthermore, the C 4 bacterium grew well, and produced a significant level of keratinase when using wool and feather as substrates. Supplementing the medium with yeast extract and peptone shortened the time required for feather degradation, but delayed the onset of the wool keratin hydrolysis with two days. The predominant amino acids released in feather hydrolysate were tyrosine, phenylalanine, and histidine. In contrast, the wool lysate was rich in aspartic acid, methionine, tyrosine, phenylalanine, histidine, and lysine. Results established that utilizing the C 4 strain for keratin degradation in waste management holds considerable potential.

Effect of pH and Temperature on Bacillus sp. R2 Chitinase Activity and Stability

Procedia Technology, 2016

Effect of Metal Ions, Chemical Agents, and Organic Solvent on Bacillus Sp.R2 Chitinase Activity

Procedia Technology, 2016

Chitinases (EC 3.2.1.14), which hydrolyze β-1,4-glucosidic bonds of chitin are widely distributed... more Chitinases (EC 3.2.1.14), which hydrolyze β-1,4-glucosidic bonds of chitin are widely distributed in the biological word and have received an increased attention during the last two decades due to their wider range of biotechnological applications specially in the agricultural, medical, industrial and environmental fields. To improve the chitinase production of Bacillus sp. R2 ,two cell immobilization strategies were conducted. The physical adsorption on different carriers showed that the highest chitinase activity was obtained by the carrier luffa pulp followed by flaked crab shell chitin with enhancement of 1.42 and 1.27 fold respectively. The entrapment immobilization in different gel matrices indicated that cell entrapment in 2% agar and 2% calcium alginate gave the highest chitinase activity 44 U/ml and 42.51 U/ml, respectively. The successive reuse of immobilized cells was investigated for 7 cycles over a period of one month. The results revealed that agar beads retained 100% or more of its original chitinase activity after 4 cycles and 86.5%, 75.5% and 60.9% of activity after the 5 th , 6 th and 7 th cycle respectively. On the other hand the alginate beads 2% and 4% retained more than 65% and 79.5% of their initial activity after 4 cycles. The effect of storage stability of beads on cell viability and chitinase production also was examined. The result showed that the longer shelf life was attained with agar beads which retain 50% of its initial activity after one month. From these optimizations, we recommend the physical adsorption immobilization on luffa pulp carrier for chitinase continuous production.

Chitinous wastes utilization and multiple enzymes production through newly isolated marine Bacillus sp.CHB

Extracellular alkaline protease was produced during aerobic cultivation of Bacillus subtilis DB10... more Extracellular alkaline protease was produced during aerobic cultivation of Bacillus subtilis DB100 (pS1) in a medium containing 2% chicken feather waste (CFW) for 48 hrs. The optimal pH and temperature of alkaline protease were 7.2 and 45°C respectively. The crude enzyme solution was precipitated with solid (NH 4 ) 2 SO 4 (65% saturation) and dialyzed against 0.1 M Tris-HCl, pH 7.5 containing 10 mM CaCl 2 . Purification was carried out using DE-52 ion exchanger column followed by gel filtration in Sephadex G-50 column. The protein content in fractions of DE-52 ion exchange & Sephadex G-50 was detected by measuring absorbance at 280 nm. The activity of alkaline protease was detected in the same fraction by its specific methods. After dialysis, about 91% of the alkaline protease total units were retained with 18.7 fold purification. After DE-52 column, about 84.6 % of the alkaline protease total units were retained and about 56 fold purification was achieved. After Sephadex G-50 column, about 68.5% of the enzyme total units were retained and about 75.5 fold purification was obtained. SDS -PAGE was carried out to check the purity of the alkaline protease enzyme sample. The resulted single protein band indicated that the enzyme was almost purified and corresponds to a molecular weight about 27 KDa.

Promissing Antifungal Chitinase from Marine Strain of Bacillus

ABSTRACT

The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poult... more The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poultry feather and sheep wool, has made keratinase production by microorganisms highly important to the biotechnological industry. A protein-degrading bacterium (C 4) was isolated from compost. Based on morphology and biochemical tests, along with 16S rRNA sequencing, the isolated C 4 was tentatively identified as Bacillus sp. C 4 (2008). The proteolytic activity of the Bacillus sp. C 4 strain was broadly specific; it degraded keratinous and non-keratinous proteins to different degrees. Pea pods as substrate generated the highest protease production, followed by soybean meal and sheep wool. Notwithstanding, using wool keratin as a sole source of carbon and nitrogen yielded the highest level of soluble proteins. Furthermore, the C 4 bacterium grew well, and produced a significant level of keratinase when using wool and feather as substrates. Supplementing the medium with yeast extract and pep...

Polish journal of microbiology / Polskie Towarzystwo Mikrobiologów = The Polish Society of Microbiologists, 2005

Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x t... more Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x to apparent homogeneity using ammonium sulfate precipitation followed by ion exchange chromatography on Q- and S-Sepharose column. Both of the purified lipases are monomeric protein with molecular weight of 15.5 and 54.97 KDa respectively. The optimal activities of the enzymes were at 45 and 50 degrees C and pHs 10.0 and 9.0. Calcium ions increase thermostability of both purified lipases I and II. The purified lipase I showed no metal ion dependence for its activity since EDTA up to 10 mM has no effect on the enzyme activity. However purified lipase II showed slight inhibition by EDTA at the same concentration. Moreover, a serine protease inhibitor, PMSF showed an inhibitory effect on both purified enzymes.

Molecular Cloning and Expression in Escherichia coli of Pseudomonas aeruginosa lipase gene

Biotechnology(Faisalabad), 2006

Cancer Cell, 2011

The aim of the present study is to evaluate the transition metals overload in Abu-Qir Bay in Egyp... more The aim of the present study is to evaluate the transition metals overload in Abu-Qir Bay in Egypt, as compared to a less polluted area (reference area) through some biomarkers of oxidative stress. Catalase enzyme activity, malondialdehyde (MDA) concentration and DNA damage (number of apurinic/apyrimidinic sites) were the tested biomarkers. The levels of iron and copper in Mugil cephalus liver tissues were significantly higher in samples from the polluted area as compared to the reference area: Fe: 407 ± 38 vs. 216 ± 21 lg/g wet wt; p = 0.008, Cu: 54 ± 6 vs. 17.7 ± 4 lg/g wet wt; p = 0.0001. This could account for the observed increase in MDA concentration (15.7 ± 5.7 vs. 2.5 ± 0.5 U/g; p = 0.035), and the elevated number of AP sites (13.9 ± 2.6 vs. 0.37 ± 0.2 AP site/1 · 10 5 bp; p = 0.0001). Similarly, the activity of catalase enzyme responsible for the cellular defense was significantly high (58.3 ± 12.2 vs. 28.4 ± 4.0 U/mg; p = 0.032). The present data indicated a clear relationship between the pollution degree of the above marine environment and both biochemical and molecular responses of the piscine system.

International Journal of Biological Chemistry, 2010

Optimizing the biodegradation of two keratinous wastes through a Bacillus subtilis recombinant strain using a response surface methodology

Biodegradation, 2010

Statistical optimization of the biodegradation of two keratinous wastes directed by Bacillus subt... more Statistical optimization of the biodegradation of two keratinous wastes directed by Bacillus subtilis recombinant cells was carried out by means of a response surface methodology. A Box-Behnken design was employed to predict the optimal levels of three variables namely, keratin percent, incubation time and inoculum size. Analysis of variance revealed that, only keratin percent had the highest significant effect. Canonical analysis and ridge max analysis were used to get the optimal levels of the three predictors along with the optimum levels of the responses. The optimal sets of predicted and validated levels of the three variables were [7.69% (w/v) feathers, 96.58 h and 1.28% (v/v) inoculum size] and [8% (w/v) feathers, 98.45 h, 3.9% (v/v) inoculum size] to achieve the highest levels of soluble proteins (1.25-1.7 mg/ml) and NH(2)-free amino groups (245.82-270.0 μmol leucine/ml), respectively upon using three optimized feathers-based media. These values represented 83.67-100% and 100% adequacy for the models of soluble proteins and NH(2)-free amino groups, respectively. While, [8.23% (w/v) sheep wool, 5.52% (v/v) inoculum size and 46.58 h] and [8.33% (w/v) sheep wool, 5.89% (v/v) inoculum size and 63.46 h] were the optimal sets of predicted and validated levels of the above variables to achieve the highest yields of soluble proteins (3.4-4.6 mg/ml) and NH(2)-free amino groups (290.9-302.0 μmol leucine/ml), respectively upon using three optimized sheep wool-based media. These values represented 100% adequacy for the models of soluble proteins and NH(2)-free amino groups. By the end of the optimization strategy, a fold enhancement (2.14-2.43 and 1.78-2.12) in the levels of released soluble proteins and NH(2)-free amino groups, respectively was obtained upon using three optimized feathers-based media. However, a fold enhancement (4.25-5.75 and 2.42-2.5) in the levels of soluble proteins and NH(2)-free amino groups, respectively was obtained upon using three optimized sheep wool-based media. Data would encourage pilot scale optimization of the biodegradation of these wastes.

Applied Biochemistry and Biotechnology, 2000

Bacillus subtilis Bios 11 strain was previously isolated and identified. This strain naturally pr... more Bacillus subtilis Bios 11 strain was previously isolated and identified. This strain naturally produces a high level of α-amylase. The multicopy (pS1) plasmid that carries the complete alkaline protease aprA gene was introduced to this host strain by transformation. The newly constructed strain was found to express the aprA gene and produces a high level of alkaline protease. The level of α-amylase production was not affected compared with the parent strain. The pS1 plasmid in the new host was proved to be segregationally and structurally stable, and the multicopy aprA gene was expressed at the stationary phase. This expression did not affect growth rate and sporulation frequency. Moreover, the level of α-amylase was maintained. Both alkaline protease and α-amylase enzymes were purified using a single-step affinity chromatography column. The use of the newly constructed strain would be valuable to the enzyme industry and would promote recycling of some food-processing wastes.

Expression of alkaline proteinase gene in two recombinant Bacillus cereus feather-degrading strains

Folia Microbiologica, 2010

Two Bacillus cereus feather-degrading strains (23/1 and 6/2) were transformed using a recombinant... more Two Bacillus cereus feather-degrading strains (23/1 and 6/2) were transformed using a recombinant plasmid p5.2 carrying the alkaline proteinase gene (aprE). A high level of the aprE gene expression was observed when the recombinant strains were grown on sporulation medium. The expression of the aprE gene proceeded during the early stationary phase and the p5.2 plasmid was segregationally and structurally stable in both strains. The two recombinant strains grown on a mineral medium with 1 % chicken feather as source of energy, carbon and nitrogen exhibited higher proteolytic activity (≈6-fold and 2.4-fold higher for strains 23/1 (p5.2) and 6/2 (p5.2), respectively. Keratinolytic activity increased ≈3.5-fold and 4.15-fold, respectively. The keratinolytic activity further increased when an optimized medium with yeast extract and corn oil was used. Considerable amounts of free amino acids were obtained after the biodegradation of feather which makes the new strains promising for application in feather-waste treatment to, e.g., transformation to animal feedstuff.

Procedia Technology, 2015

A simple assay for determining activities of phosphopentomutase from a hyperthermophilic bacterium Thermotoga maritima

Analytical biochemistry, Jan 15, 2016

Phosphopentomutase (PPM) catalyzes the interconversion of α-D-(deoxy)-ribose 1-phosphate and α-D-... more Phosphopentomutase (PPM) catalyzes the interconversion of α-D-(deoxy)-ribose 1-phosphate and α-D-(deoxy)-ribose 5-phosphate. We developed a coupled or uncoupled enzymatic assay with an enzyme nucleoside phosphorylase for determining PPM activities on D-ribose 5-phosphate at a broad temperature range from 30 to 90 °C. This assay not only is simple and highly sensitive but also does not require any costly special instrument. Via this technology, an open reading frame TM0167 from a thermophilic bacterium Thermotoga maritima putatively encoding PPM was cloned. The recombinant PPM was overexpressed in Escherichia coli Rosetta. This enzyme has the highest activity at 90 °C. MnCl2 (0.1 mM) and 50 μM α-D-glucose 1,6-bisphosphate are cofactors. The kinetic parameters of Km and kcat are 1.2 mM and 185 s(-1) at 90 °C, respectively. The enzyme has a half-life time of up to 156 min at 90 °C. This enzyme is the most active and thermostable PPM reported to date.

Molecular Cloning and Expression in Escherichia coli of Pseudomonas aeruginosa lipase gene

Biotechnology, 2006

Polish Journal of Microbiology Polskie Towarzystwo Mikrobiologow the Polish Society of Microbiologists, Feb 1, 2005

Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x t... more Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x to apparent homogeneity using ammonium sulfate precipitation followed by ion exchange chromatography on Q-and S-Sepharose column. Both of the purified lipases are monomeric protein with molecular weight of 15.5 and 54.97 KDa respectively. The optimal activities of the enzymes were at 45 and 50°C and pHs 10.0 and 9.0. Calcium ions increase thermostability of both purified lipases I and II. The purified lipase I showed no metal ion dependence for its activity since EDTA up to 10 mM has no effect on the enzyme activity. However purified lipase II showed slight inhibition by EDTA at the same concentration. Moreover, a serine protease inhibitor, PMSF showed an inhibitory effect on both purified enzymes. K e y w o r d s: Lipases, Pseudomonas aeruginosa Ps-x, 16S rRNA Abbreviations: SDS-PAGE; sodium dodecyl sulfate polyacrylamide gel electrophoresis, Q-Sepharose; Quaternary amino methyl Sepharose, EDTA; ethylene diamine tetraacetic acid.

The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poult... more The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poultry feather and sheep wool, has made keratinase production by microorganisms highly important to the biotechnological industry. A proteindegrading bacterium (C 4 ) was isolated from compost. Based on morphology and biochemical tests, along with 16S rRNA sequencing, the isolated C 4 was tentatively identified as Bacillus sp. C 4 (2008). The proteolytic activity of the Bacillus sp. C 4 strain was broadly specific; it degraded keratinous and non-keratinous proteins to different degrees. Pea pods as substrate generated the highest protease production, followed by soybean meal and sheep wool. Notwithstanding, using wool keratin as a sole source of carbon and nitrogen yielded the highest level of soluble proteins. Furthermore, the C 4 bacterium grew well, and produced a significant level of keratinase when using wool and feather as substrates. Supplementing the medium with yeast extract and peptone shortened the time required for feather degradation, but delayed the onset of the wool keratin hydrolysis with two days. The predominant amino acids released in feather hydrolysate were tyrosine, phenylalanine, and histidine. In contrast, the wool lysate was rich in aspartic acid, methionine, tyrosine, phenylalanine, histidine, and lysine. Results established that utilizing the C 4 strain for keratin degradation in waste management holds considerable potential.

Effect of pH and Temperature on Bacillus sp. R2 Chitinase Activity and Stability

Procedia Technology, 2016

Effect of Metal Ions, Chemical Agents, and Organic Solvent on Bacillus Sp.R2 Chitinase Activity

Procedia Technology, 2016

Chitinases (EC 3.2.1.14), which hydrolyze β-1,4-glucosidic bonds of chitin are widely distributed... more Chitinases (EC 3.2.1.14), which hydrolyze β-1,4-glucosidic bonds of chitin are widely distributed in the biological word and have received an increased attention during the last two decades due to their wider range of biotechnological applications specially in the agricultural, medical, industrial and environmental fields. To improve the chitinase production of Bacillus sp. R2 ,two cell immobilization strategies were conducted. The physical adsorption on different carriers showed that the highest chitinase activity was obtained by the carrier luffa pulp followed by flaked crab shell chitin with enhancement of 1.42 and 1.27 fold respectively. The entrapment immobilization in different gel matrices indicated that cell entrapment in 2% agar and 2% calcium alginate gave the highest chitinase activity 44 U/ml and 42.51 U/ml, respectively. The successive reuse of immobilized cells was investigated for 7 cycles over a period of one month. The results revealed that agar beads retained 100% or more of its original chitinase activity after 4 cycles and 86.5%, 75.5% and 60.9% of activity after the 5 th , 6 th and 7 th cycle respectively. On the other hand the alginate beads 2% and 4% retained more than 65% and 79.5% of their initial activity after 4 cycles. The effect of storage stability of beads on cell viability and chitinase production also was examined. The result showed that the longer shelf life was attained with agar beads which retain 50% of its initial activity after one month. From these optimizations, we recommend the physical adsorption immobilization on luffa pulp carrier for chitinase continuous production.

Chitinous wastes utilization and multiple enzymes production through newly isolated marine Bacillus sp.CHB

Extracellular alkaline protease was produced during aerobic cultivation of Bacillus subtilis DB10... more Extracellular alkaline protease was produced during aerobic cultivation of Bacillus subtilis DB100 (pS1) in a medium containing 2% chicken feather waste (CFW) for 48 hrs. The optimal pH and temperature of alkaline protease were 7.2 and 45°C respectively. The crude enzyme solution was precipitated with solid (NH 4 ) 2 SO 4 (65% saturation) and dialyzed against 0.1 M Tris-HCl, pH 7.5 containing 10 mM CaCl 2 . Purification was carried out using DE-52 ion exchanger column followed by gel filtration in Sephadex G-50 column. The protein content in fractions of DE-52 ion exchange & Sephadex G-50 was detected by measuring absorbance at 280 nm. The activity of alkaline protease was detected in the same fraction by its specific methods. After dialysis, about 91% of the alkaline protease total units were retained with 18.7 fold purification. After DE-52 column, about 84.6 % of the alkaline protease total units were retained and about 56 fold purification was achieved. After Sephadex G-50 column, about 68.5% of the enzyme total units were retained and about 75.5 fold purification was obtained. SDS -PAGE was carried out to check the purity of the alkaline protease enzyme sample. The resulted single protein band indicated that the enzyme was almost purified and corresponds to a molecular weight about 27 KDa.

Promissing Antifungal Chitinase from Marine Strain of Bacillus

ABSTRACT

The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poult... more The ability to hydrolyze keratin, a rigid and strongly cross-linked protein in the waste of poultry feather and sheep wool, has made keratinase production by microorganisms highly important to the biotechnological industry. A protein-degrading bacterium (C 4) was isolated from compost. Based on morphology and biochemical tests, along with 16S rRNA sequencing, the isolated C 4 was tentatively identified as Bacillus sp. C 4 (2008). The proteolytic activity of the Bacillus sp. C 4 strain was broadly specific; it degraded keratinous and non-keratinous proteins to different degrees. Pea pods as substrate generated the highest protease production, followed by soybean meal and sheep wool. Notwithstanding, using wool keratin as a sole source of carbon and nitrogen yielded the highest level of soluble proteins. Furthermore, the C 4 bacterium grew well, and produced a significant level of keratinase when using wool and feather as substrates. Supplementing the medium with yeast extract and pep...

Polish journal of microbiology / Polskie Towarzystwo Mikrobiologów = The Polish Society of Microbiologists, 2005

Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x t... more Two different extracellular lipases were isolated and purified from Pseudomonas aeruginosa Ps-x to apparent homogeneity using ammonium sulfate precipitation followed by ion exchange chromatography on Q- and S-Sepharose column. Both of the purified lipases are monomeric protein with molecular weight of 15.5 and 54.97 KDa respectively. The optimal activities of the enzymes were at 45 and 50 degrees C and pHs 10.0 and 9.0. Calcium ions increase thermostability of both purified lipases I and II. The purified lipase I showed no metal ion dependence for its activity since EDTA up to 10 mM has no effect on the enzyme activity. However purified lipase II showed slight inhibition by EDTA at the same concentration. Moreover, a serine protease inhibitor, PMSF showed an inhibitory effect on both purified enzymes.

Molecular Cloning and Expression in Escherichia coli of Pseudomonas aeruginosa lipase gene

Biotechnology(Faisalabad), 2006

Cancer Cell, 2011

The aim of the present study is to evaluate the transition metals overload in Abu-Qir Bay in Egyp... more The aim of the present study is to evaluate the transition metals overload in Abu-Qir Bay in Egypt, as compared to a less polluted area (reference area) through some biomarkers of oxidative stress. Catalase enzyme activity, malondialdehyde (MDA) concentration and DNA damage (number of apurinic/apyrimidinic sites) were the tested biomarkers. The levels of iron and copper in Mugil cephalus liver tissues were significantly higher in samples from the polluted area as compared to the reference area: Fe: 407 ± 38 vs. 216 ± 21 lg/g wet wt; p = 0.008, Cu: 54 ± 6 vs. 17.7 ± 4 lg/g wet wt; p = 0.0001. This could account for the observed increase in MDA concentration (15.7 ± 5.7 vs. 2.5 ± 0.5 U/g; p = 0.035), and the elevated number of AP sites (13.9 ± 2.6 vs. 0.37 ± 0.2 AP site/1 · 10 5 bp; p = 0.0001). Similarly, the activity of catalase enzyme responsible for the cellular defense was significantly high (58.3 ± 12.2 vs. 28.4 ± 4.0 U/mg; p = 0.032). The present data indicated a clear relationship between the pollution degree of the above marine environment and both biochemical and molecular responses of the piscine system.

International Journal of Biological Chemistry, 2010

Optimizing the biodegradation of two keratinous wastes through a Bacillus subtilis recombinant strain using a response surface methodology

Biodegradation, 2010

Statistical optimization of the biodegradation of two keratinous wastes directed by Bacillus subt... more Statistical optimization of the biodegradation of two keratinous wastes directed by Bacillus subtilis recombinant cells was carried out by means of a response surface methodology. A Box-Behnken design was employed to predict the optimal levels of three variables namely, keratin percent, incubation time and inoculum size. Analysis of variance revealed that, only keratin percent had the highest significant effect. Canonical analysis and ridge max analysis were used to get the optimal levels of the three predictors along with the optimum levels of the responses. The optimal sets of predicted and validated levels of the three variables were [7.69% (w/v) feathers, 96.58 h and 1.28% (v/v) inoculum size] and [8% (w/v) feathers, 98.45 h, 3.9% (v/v) inoculum size] to achieve the highest levels of soluble proteins (1.25-1.7 mg/ml) and NH(2)-free amino groups (245.82-270.0 μmol leucine/ml), respectively upon using three optimized feathers-based media. These values represented 83.67-100% and 100% adequacy for the models of soluble proteins and NH(2)-free amino groups, respectively. While, [8.23% (w/v) sheep wool, 5.52% (v/v) inoculum size and 46.58 h] and [8.33% (w/v) sheep wool, 5.89% (v/v) inoculum size and 63.46 h] were the optimal sets of predicted and validated levels of the above variables to achieve the highest yields of soluble proteins (3.4-4.6 mg/ml) and NH(2)-free amino groups (290.9-302.0 μmol leucine/ml), respectively upon using three optimized sheep wool-based media. These values represented 100% adequacy for the models of soluble proteins and NH(2)-free amino groups. By the end of the optimization strategy, a fold enhancement (2.14-2.43 and 1.78-2.12) in the levels of released soluble proteins and NH(2)-free amino groups, respectively was obtained upon using three optimized feathers-based media. However, a fold enhancement (4.25-5.75 and 2.42-2.5) in the levels of soluble proteins and NH(2)-free amino groups, respectively was obtained upon using three optimized sheep wool-based media. Data would encourage pilot scale optimization of the biodegradation of these wastes.

Applied Biochemistry and Biotechnology, 2000

Bacillus subtilis Bios 11 strain was previously isolated and identified. This strain naturally pr... more Bacillus subtilis Bios 11 strain was previously isolated and identified. This strain naturally produces a high level of α-amylase. The multicopy (pS1) plasmid that carries the complete alkaline protease aprA gene was introduced to this host strain by transformation. The newly constructed strain was found to express the aprA gene and produces a high level of alkaline protease. The level of α-amylase production was not affected compared with the parent strain. The pS1 plasmid in the new host was proved to be segregationally and structurally stable, and the multicopy aprA gene was expressed at the stationary phase. This expression did not affect growth rate and sporulation frequency. Moreover, the level of α-amylase was maintained. Both alkaline protease and α-amylase enzymes were purified using a single-step affinity chromatography column. The use of the newly constructed strain would be valuable to the enzyme industry and would promote recycling of some food-processing wastes.

Expression of alkaline proteinase gene in two recombinant Bacillus cereus feather-degrading strains

Folia Microbiologica, 2010

Two Bacillus cereus feather-degrading strains (23/1 and 6/2) were transformed using a recombinant... more Two Bacillus cereus feather-degrading strains (23/1 and 6/2) were transformed using a recombinant plasmid p5.2 carrying the alkaline proteinase gene (aprE). A high level of the aprE gene expression was observed when the recombinant strains were grown on sporulation medium. The expression of the aprE gene proceeded during the early stationary phase and the p5.2 plasmid was segregationally and structurally stable in both strains. The two recombinant strains grown on a mineral medium with 1 % chicken feather as source of energy, carbon and nitrogen exhibited higher proteolytic activity (≈6-fold and 2.4-fold higher for strains 23/1 (p5.2) and 6/2 (p5.2), respectively. Keratinolytic activity increased ≈3.5-fold and 4.15-fold, respectively. The keratinolytic activity further increased when an optimized medium with yeast extract and corn oil was used. Considerable amounts of free amino acids were obtained after the biodegradation of feather which makes the new strains promising for application in feather-waste treatment to, e.g., transformation to animal feedstuff.