Based on garden cress significantly used for phytoremediation, the antioxidant system included an... more Based on garden cress significantly used for phytoremediation, the antioxidant system included antioxidant-phenolic compounds and antioxidant-enzymes of 6-day-garden cress sprouts (GCS) were assessed as potential bio-indicators for cadmium (Cd) and lead (Pb) contamination. Total phenolic and flavonoid contents of GCS germinated under Cd and Pb treatments (25–150 mg kg−1) gradually increased with increasing concentration of metals and peaked by 2.0, 2.6, and 2.5, 2.3 folds at 150 mg kg−1, respectively. By using DPPH, ABTS, and PMC antioxidant assays, the total antioxidant activity of phenolic compounds of GCS increased 6.1, 13.0, and 5.8-fold for Cd and 5.9, 14.6, and 8.2-fold for Pb at 150 mg kg−1, respectively. The antioxidant enzymes of GCS (POD, CAT, GR, and GST) were significantly activated in response to Cd and Pb stress, and two new electrophoretic POD bands were detected. GCS was absorbed 19.0% and 21.3% of Cd and Pb at 150 mg metal kg−1, respectively. In conclusion, the appr...
Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present ... more Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present study, Sarcocystis fusiformis was recognized in Egyptian water buffaloes based on histological observation and molecular analysis of internal transcribed spacer 1( ITS1), 18S ribosomal RNA (18S rRNA) and cytochrome c oxidase subunit I (COX-1) gene fragments. Chemotherapy and vaccines against Sarcocystis spp. could potentially target proteases because they may play a crucial role in the infection. Cysteine proteases are multifunctional enzymes involved in vital metabolic processes. However, the involvement of proteases in S. fusiform infection has not yet been characterized. Here, the purification and study on some biochemical properties of protease isolated from cysts of S. fusiform were carried out. Protease with a molecular weight of 100 kDa was purified. The purified protease exhibited maximum activity at pH 6 and a temperature of 50°C. The Michaelis-Menten constant (Km), the maximum ...
Saw palmetto seeds (SPS) contain essential phenolic compounds that provide antioxidant, antimicro... more Saw palmetto seeds (SPS) contain essential phenolic compounds that provide antioxidant, antimicrobial, anti-inflammatory, and anti-diabetic benefits when added to food. Maximized/improved production of these valuable phenolic compounds is the main purpose of this study. Solid-state fermentation (SSF) is a promising processing technique that positively alters the levels of health-promoting compounds in plants and plant residues. Here, a central composite design matrix (16 runs) and response surface methodology were experimentally applied to investigate the best SSF conditions and their interactions for maximum production of phenolic compounds from SPS. A good correlation between actual and expected results was observed with higher multiple coefficients (R2 ~ 0.93–0.97) and strongly significant P values (< 0.0001) proving the accuracy of the statistical model/design. Under optimized SSF conditions, temperature 30 °C, moisture 10%, pH 7.0, and fermentation time 6 days, the total phe...
The study aims to evaluate the relation between peroxidases of day-6 garden cress sprouts and phe... more The study aims to evaluate the relation between peroxidases of day-6 garden cress sprouts and phenolic compounds. Three cationic, three anionic, and two unbounded peroxidases were separated from day-6 garden cress sprouts. Cationic (GCP1) and anionic (GCP2) peroxidases were purified with molecular masses of 25 and 40 kDa, respectively. The Km values of GCP1 toward H2 O2 and guaiacol were lower than GCP2. The anionic GCP2 exhibited high affinity toward some lignin monomers, sinapyl alcohol, coniferyl alcohol, cinnamic and ferulic acids. Therefore, GCP2 is considered as a lignin peroxidase and contributed in lignin synthesis. The activity of GCP1 and GCP2 was stable at a wide pH range 5.5-8.0 and 6.0-7.5, respectively. Both peroxidases showed the same thermal stability range 20-50°C. GCP2 was more resistant against the effect of metal ions than GCP1. GCP2 showed high ability to remove of phenol and p-chlorophenol from effluent compared to GCP1. PRACTICAL APPLICATIONS: Generally, garde...
The obtained garden cress 6-day sprouts phenolic-rich extract (GCSP) contained efficient health-p... more The obtained garden cress 6-day sprouts phenolic-rich extract (GCSP) contained efficient health-promoting antioxidant-phenolic compounds. To improve the stability, bioavailability, and functional properties of these valuable phenolic compounds, GCSP was encapsulated by freeze-drying technique using different ratios of garden cress gum (GG) and maltodextrin (M) in the absence and presence of sonication (S). The prepared S/GG-microcapsule retained the highest phenolic content (95%), antioxidant activity (141.6%), and encapsulation efficiency (98.2%). It displayed the highest bio-accessibility of GCSP-phenolic compounds in simulated intestine fluid (87%) and demonstrated the greatest storage-stability at 40 °C for 60 days. S/GG-microcapsule possessed better physical properties including moisture, solubility, swelling, and morphological structures using SEM. The main spectral features, crosslinking, and improved thermal stability were demonstrated for S/GG-microcapsule using FTIR and th...
Chia seeds (CS) are becoming increasingly consumed due to their great nutritional and therapeutic... more Chia seeds (CS) are becoming increasingly consumed due to their great nutritional and therapeutic properties. In this study, solid-state fermentation (SSF) of CS by Trichoderma reesei was employed to maximize the production of the antioxidant-phenolic compounds and some fungal phenolic-associated enzymes (α-amylase, xylanase, β-glucosidase, polygalacturonase, and phenylalanine ammonia-lyase). The SSF-conditions were statistically optimized using response surface methodology (RSM). In the statistical model, four variables were analyzed at two levels. According to RSM, the adjusted R2 (< 0.9) is reasonably consistent with the predicted R2 (< 0.9), indicating that the statistical model is valid. The optimal conditions for maximum production of both phenolic compounds and fungal phenolic-associated enzymes were found to be 28 °C, pH 7.0, 20% moisture, and 7-day fermentation. The total phenolic content of fermented CS (FCS) increased 23 folds and total antioxidant activity was enha...
Abstract: Medicinal therapy requires careful assessment of effective treatment offering an accept... more Abstract: Medicinal therapy requires careful assessment of effective treatment offering an acceptable safety over human health. This study aimed to evaluate the influence of walnuts diet on streptozotocin-induced diabetic mice. Comprehen-sive systems of biochemical markers and genomic screening were used to achieve this task. Experimental animals were divided into four groups (negative and positive control, diabetic group and diabetic group fed on walnuts diet). Diabetes was induced by i.p. injection of streptozotocin (STZ) for 3 successive days. Biomarkers assay, gene expression analysis, comet assay, quantitation of DNA fragmentation and micronucleus assay were investigated. The results suggested that walnuts could act as diabetic therapy without adverse effects at metabolic activity and molecular levels in diabetic mice.
Hepatitis C virus (HCV) is the leading cause of liver diseases worldwide. At present, drug combin... more Hepatitis C virus (HCV) is the leading cause of liver diseases worldwide. At present, drug combinations of different classes of direct-acting antiviral agents (DAAs) are used as treatment options for HCV, in which sofosbuvir (SOF) is the common DAA among different therapeutic regimes. In Egypt, SOF plus daclatasvir (DCV) is the widely used anti-HCV treatment protocol. Herein, we aimed to assess the association between 3 single-nucleotide polymorphisms (SNPs) at the genes coding for 2 SOF metabolizing enzymes: histidine triad nucleotide-binding protein 1 (HINT1) rs4696/rs7728773 and nucleoside diphosphate kinase 1 (NME1) rs3760468, together with the most potent anti-HCV innate molecule, i.e., Interferon lambda 3 (IFNL3) rs12979860 and the response to SOF/DCV in Egyptian patients chronically infected with genotype 4 (GT4). SNPs were genotyped using real-time PCR in DNA from patients who achieved sustained virological response (SVR) at 12 weeks post-SOF/DCV treatment (i.e., responders; n = 188) and compared with patients who failed to achieve SVR12 (i.e., non-responders; n = 109) or healthy controls (n = 62). Our results demonstrated that patients bearing HINT1 rs7728773 CT/TT (odds ratio 2.119, 95% CI 1.263-3.559, p = 0.005) and IFNL3 rs12979860 CC (odds ratio 3.995, 95% CI 2.126-7.740, p = 0.0001) were more likely to achieve SVR12. However, neither HINT1 rs4696 nor NME1 rs3760468 seems to contribute to the responsiveness to SOF/DCV. Binary regression analysis defined 5 predictor factors independently associated with SVR12: age, bilirubin, HB, early stages of fibrosis, and combined HINT1 rs7728773 and IFNL3 rs12979860 favorable and mixed genotypes (odds ratio 3.134, 95% CI 1.518-6.47, p = 0.002), and that was confirmed by the combined ROC curve for the 5 predictor factors (AUC = 0.91, 95% CI 0.869-0.95, P = 0.0001). In conclusion, these data suggest that the two SNPs have the potential in predicting the response rate to SOF/DCV treatment in patients infected with HCV GT4. This study is the first to investigate the pharmacogenetics of SOF metabolizing enzyme and introduce HINT1 rs7728773 as a novel SNP that predict the treatment efficacy.
Production of amylases by fungi under solid-state fermentation is considered the best methodology... more Production of amylases by fungi under solid-state fermentation is considered the best methodology for commercial scaling that addresses the ever-escalating needs of the worldwide enzyme market. Here response surface methodology (RSM) was used for the optimization of process variables for α-amylase enzyme production from Trichoderma virens using watermelon rinds (WMR) under solid-state fermentation (SSF). The statistical model included four variables, each detected at two levels, followed by model development with partial purification and characterization of α-amylase. The partially purified α-amylase was characterized with regard to optimum pH, temperature, kinetic constant, and substrate specificity. The results indicated that both pH and moisture content had a significant effect (P < 0.05) on α-amylase production (880 U/g) under optimized process conditions at a 3-day incubation time, moisture content of 50%, 30 °C, and pH 6.98. Statistical optimization using RSM showed R2 values of 0.9934, demonstrating the validity of the model. Five α-amylases were separated by using DEAE-Sepharose and characterized with a wide range of optimized pH values (pH 4.5-9.0), temperature optima (40-60 °C), low Km values (2.27-3.3 mg/mL), and high substrate specificity toward large substrates. In conclusion, this study presents an efficient and green approach for utilization of agro-waste for production of the valuable α-amylase enzyme using RSM under SSF. RSM was particularly beneficial for the optimization and analysis of the effective process parameters.
Xylan saccharification is a key step in many important biotechnological applications. Xylose is t... more Xylan saccharification is a key step in many important biotechnological applications. Xylose is the main product of xylan degradation and is a major xylanase inhibitor in a bioreactor; however, xylose-binding site of xylanase is not discovered yet. Evolving of xylose-tolerant xylanase variants will reduce the cost of xylanases in industry. Glycoside hydrolase family-10 thermostable Geobacillus stearothermophilus xylanase XT6 is non-competitively inhibited by xylose with inhibition constant ki equals to 12.2 mM. In the absence of X-ray crystallography of xylanase-xylose complex, unbiased random mutagenesis of the whole xylanase gene was done by error-prone polymerase chain reaction constructing a huge library. Screening a part of the library revealed xylose-tolerant mutants having three mutations, M116I, L131P and L133V, clustered in the N-terminus of α-helix 3. The best xylose-tolerant mutant showed higher ki and catalytic capability than that of the parent by 3.5- and 3-fold, respectively. In addition, kcat increased 4.5-fold and KM decreased 2-fold. The molecular docking of xylose into xylanase XT6 structure showed that xylose binds into a small pocket between N-terminus of α-helices 3 and 4 and close to the three mutations. Mobility of α-helices 3 and 4, which controls catalysis rate, is restricted by xylose binding and increased by these mutations.
Planarians have been long known for their regenerative ability, which hinges on pluripotency. Rec... more Planarians have been long known for their regenerative ability, which hinges on pluripotency. Recently, however, the planarian model has been successfully established for routine toxicological screens aimed to assess overproliferation, mutagenicity and tumorigenesis. In this study, we focused on planarian tumor suppressor genes (TSGs) and their role during chemically induced carcinogenic stress in Combining and proteomic screens with exposure to human carcinogen type 1A agent cadmium (Cd), we showed that many TSGs have a function in stem cells and that, in general, exposure to Cd accelerated the onset and increased the severity of the observed phenotype. This suggested that the interaction between environmental and genetic factors plays an important role in tumor development in Therefore, we further focused on the synergistic effects of Cd exposure and knockdown (KD) at the cellular and molecular levels. Cd also produced a specific proteomic landscape in homeostatic animals, with 17...
Heat-shock protein 110 (HSP110) family members act as nucleotide exchange factors (NEF) of mammal... more Heat-shock protein 110 (HSP110) family members act as nucleotide exchange factors (NEF) of mammalian and yeast HSP70 chaperones during the ATP hydrolysis cycle. In this study, we describe the expression pattern of murine HSPA4, a member of the HSP110 family, during testis development and the consequence of HSPA4 deficiency on male fertility. HSPA4 is ubiquitously expressed in all the examined tissues. During prenatal and postnatal development of gonad, HSPA4 is expressed in both somatic and germ cells; however, expression was much higher in germ cells of prenatal gonads. Analyses ofHspa4-deficient mice revealed that all homozygous mice on the hybrid C57BL/6J×129/Sv genetic background were apparently healthy. Although HSPA4 is expressed as early as E13.5 in male gonad, a lack of histological differences betweenHspa4−/−and control littermates suggests thatHspa4deficiency does not impair the gonocytes or their development to spermatogonia. Remarkably, an increased number of theHspa4-de...
Journal of Molecular and Cellular Cardiology, 2012
Failure of molecular chaperones to direct the correct folding of newly synthesized proteins leads... more Failure of molecular chaperones to direct the correct folding of newly synthesized proteins leads to the accumulation of misfolded proteins in cells. HSPA4 is a member of the heat shock protein 110 family (HSP110) that acts as a nucleotide exchange factor of HSP70 chaperones. We found that the expression of HSPA4 is upregulated in murine hearts subjected to pressure overload and in failing human hearts. To investigate the cardiac function of HSPA4, Hspa4 knockout (KO) mice were generated and exhibited cardiac hypertrophy and fibrosis. Hspa4 KO hearts were characterized by a significant increase in heart weight/body weight ratio, elevated expression of hypertrophic and fibrotic gene markers, and concentric hypertrophy with preserved contractile function. In response to pressure overload, cardiac hypertrophy and remodeling were further aggravated in the Hspa4 KO compared to wild type (WT) mice. Cardiac hypertrophy in Hspa4 KO hearts was associated with enhanced activation of gp130-STAT3, CaMKII, and calcineurin-NFAT signaling. Protein blot and immunofluorescent analyses showed a significant accumulation of polyubiquitinated proteins in cardiac cells of Hspa4 KO mice. These results suggest that the myocardial remodeling of Hspa4 KO mice is due to accumulation of misfolded proteins resulting from impaired chaperone activity. Further analyses revealed a significant increase in cross sectional area of cardiomyocytes, and in expression levels of hypertrophic markers in cultured neonatal Hspa4 KO cardiomyocytes suggesting that the hypertrophy of mutant mice was a result of primary defects in cardiomyocytes. Gene expression profile in hearts of 3.5-week-old mice revealed a differentially expressed gene sets related to ion channels, muscle-specific contractile proteins and stress response. Taken together, our in vivo data demonstrate that Hspa4 gene ablation results in cardiac hypertrophy and fibrosis, possibly, through its role in protein quality control mechanism.
Background Molecular chaperones assist protein folding, facilitate degradation of misfolded polyp... more Background Molecular chaperones assist protein folding, facilitate degradation of misfolded polypeptides, and thereby maintain protein homeostasis. Impaired chaperone activity leads to defective protein quality control that is implicated in multiple skeletal muscle diseases. The heat shock protein A4 (HSPA4) acts as a co-chaperone for HSP70. Previously, we showed that Hspa4 deletion causes impaired protein homeostasis in the heart. However, its functional role in skeletal muscle has not been explored. Methods We performed a comparative phenotypic and biochemical analyses of Hspa4 knockout (KO) mice with wild-type (WT) littermates. Results HSPA4 is markedly upregulated in regenerating WT muscle in vivo, and in differentiated myoblasts in vitro. Hspa4-KO mice are marked by growth retardation and increased variability in body weight, accompanied by 35% mortality rates during the peri-weaning period. The surviving Hspa4-KO mice experienced progressive skeletal muscle myopathy, character...
Based on garden cress significantly used for phytoremediation, the antioxidant system included an... more Based on garden cress significantly used for phytoremediation, the antioxidant system included antioxidant-phenolic compounds and antioxidant-enzymes of 6-day-garden cress sprouts (GCS) were assessed as potential bio-indicators for cadmium (Cd) and lead (Pb) contamination. Total phenolic and flavonoid contents of GCS germinated under Cd and Pb treatments (25–150 mg kg−1) gradually increased with increasing concentration of metals and peaked by 2.0, 2.6, and 2.5, 2.3 folds at 150 mg kg−1, respectively. By using DPPH, ABTS, and PMC antioxidant assays, the total antioxidant activity of phenolic compounds of GCS increased 6.1, 13.0, and 5.8-fold for Cd and 5.9, 14.6, and 8.2-fold for Pb at 150 mg kg−1, respectively. The antioxidant enzymes of GCS (POD, CAT, GR, and GST) were significantly activated in response to Cd and Pb stress, and two new electrophoretic POD bands were detected. GCS was absorbed 19.0% and 21.3% of Cd and Pb at 150 mg metal kg−1, respectively. In conclusion, the appr...
Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present ... more Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present study, Sarcocystis fusiformis was recognized in Egyptian water buffaloes based on histological observation and molecular analysis of internal transcribed spacer 1( ITS1), 18S ribosomal RNA (18S rRNA) and cytochrome c oxidase subunit I (COX-1) gene fragments. Chemotherapy and vaccines against Sarcocystis spp. could potentially target proteases because they may play a crucial role in the infection. Cysteine proteases are multifunctional enzymes involved in vital metabolic processes. However, the involvement of proteases in S. fusiform infection has not yet been characterized. Here, the purification and study on some biochemical properties of protease isolated from cysts of S. fusiform were carried out. Protease with a molecular weight of 100 kDa was purified. The purified protease exhibited maximum activity at pH 6 and a temperature of 50°C. The Michaelis-Menten constant (Km), the maximum ...
Saw palmetto seeds (SPS) contain essential phenolic compounds that provide antioxidant, antimicro... more Saw palmetto seeds (SPS) contain essential phenolic compounds that provide antioxidant, antimicrobial, anti-inflammatory, and anti-diabetic benefits when added to food. Maximized/improved production of these valuable phenolic compounds is the main purpose of this study. Solid-state fermentation (SSF) is a promising processing technique that positively alters the levels of health-promoting compounds in plants and plant residues. Here, a central composite design matrix (16 runs) and response surface methodology were experimentally applied to investigate the best SSF conditions and their interactions for maximum production of phenolic compounds from SPS. A good correlation between actual and expected results was observed with higher multiple coefficients (R2 ~ 0.93–0.97) and strongly significant P values (< 0.0001) proving the accuracy of the statistical model/design. Under optimized SSF conditions, temperature 30 °C, moisture 10%, pH 7.0, and fermentation time 6 days, the total phe...
The study aims to evaluate the relation between peroxidases of day-6 garden cress sprouts and phe... more The study aims to evaluate the relation between peroxidases of day-6 garden cress sprouts and phenolic compounds. Three cationic, three anionic, and two unbounded peroxidases were separated from day-6 garden cress sprouts. Cationic (GCP1) and anionic (GCP2) peroxidases were purified with molecular masses of 25 and 40 kDa, respectively. The Km values of GCP1 toward H2 O2 and guaiacol were lower than GCP2. The anionic GCP2 exhibited high affinity toward some lignin monomers, sinapyl alcohol, coniferyl alcohol, cinnamic and ferulic acids. Therefore, GCP2 is considered as a lignin peroxidase and contributed in lignin synthesis. The activity of GCP1 and GCP2 was stable at a wide pH range 5.5-8.0 and 6.0-7.5, respectively. Both peroxidases showed the same thermal stability range 20-50°C. GCP2 was more resistant against the effect of metal ions than GCP1. GCP2 showed high ability to remove of phenol and p-chlorophenol from effluent compared to GCP1. PRACTICAL APPLICATIONS: Generally, garde...
The obtained garden cress 6-day sprouts phenolic-rich extract (GCSP) contained efficient health-p... more The obtained garden cress 6-day sprouts phenolic-rich extract (GCSP) contained efficient health-promoting antioxidant-phenolic compounds. To improve the stability, bioavailability, and functional properties of these valuable phenolic compounds, GCSP was encapsulated by freeze-drying technique using different ratios of garden cress gum (GG) and maltodextrin (M) in the absence and presence of sonication (S). The prepared S/GG-microcapsule retained the highest phenolic content (95%), antioxidant activity (141.6%), and encapsulation efficiency (98.2%). It displayed the highest bio-accessibility of GCSP-phenolic compounds in simulated intestine fluid (87%) and demonstrated the greatest storage-stability at 40 °C for 60 days. S/GG-microcapsule possessed better physical properties including moisture, solubility, swelling, and morphological structures using SEM. The main spectral features, crosslinking, and improved thermal stability were demonstrated for S/GG-microcapsule using FTIR and th...
Chia seeds (CS) are becoming increasingly consumed due to their great nutritional and therapeutic... more Chia seeds (CS) are becoming increasingly consumed due to their great nutritional and therapeutic properties. In this study, solid-state fermentation (SSF) of CS by Trichoderma reesei was employed to maximize the production of the antioxidant-phenolic compounds and some fungal phenolic-associated enzymes (α-amylase, xylanase, β-glucosidase, polygalacturonase, and phenylalanine ammonia-lyase). The SSF-conditions were statistically optimized using response surface methodology (RSM). In the statistical model, four variables were analyzed at two levels. According to RSM, the adjusted R2 (< 0.9) is reasonably consistent with the predicted R2 (< 0.9), indicating that the statistical model is valid. The optimal conditions for maximum production of both phenolic compounds and fungal phenolic-associated enzymes were found to be 28 °C, pH 7.0, 20% moisture, and 7-day fermentation. The total phenolic content of fermented CS (FCS) increased 23 folds and total antioxidant activity was enha...
Abstract: Medicinal therapy requires careful assessment of effective treatment offering an accept... more Abstract: Medicinal therapy requires careful assessment of effective treatment offering an acceptable safety over human health. This study aimed to evaluate the influence of walnuts diet on streptozotocin-induced diabetic mice. Comprehen-sive systems of biochemical markers and genomic screening were used to achieve this task. Experimental animals were divided into four groups (negative and positive control, diabetic group and diabetic group fed on walnuts diet). Diabetes was induced by i.p. injection of streptozotocin (STZ) for 3 successive days. Biomarkers assay, gene expression analysis, comet assay, quantitation of DNA fragmentation and micronucleus assay were investigated. The results suggested that walnuts could act as diabetic therapy without adverse effects at metabolic activity and molecular levels in diabetic mice.
Hepatitis C virus (HCV) is the leading cause of liver diseases worldwide. At present, drug combin... more Hepatitis C virus (HCV) is the leading cause of liver diseases worldwide. At present, drug combinations of different classes of direct-acting antiviral agents (DAAs) are used as treatment options for HCV, in which sofosbuvir (SOF) is the common DAA among different therapeutic regimes. In Egypt, SOF plus daclatasvir (DCV) is the widely used anti-HCV treatment protocol. Herein, we aimed to assess the association between 3 single-nucleotide polymorphisms (SNPs) at the genes coding for 2 SOF metabolizing enzymes: histidine triad nucleotide-binding protein 1 (HINT1) rs4696/rs7728773 and nucleoside diphosphate kinase 1 (NME1) rs3760468, together with the most potent anti-HCV innate molecule, i.e., Interferon lambda 3 (IFNL3) rs12979860 and the response to SOF/DCV in Egyptian patients chronically infected with genotype 4 (GT4). SNPs were genotyped using real-time PCR in DNA from patients who achieved sustained virological response (SVR) at 12 weeks post-SOF/DCV treatment (i.e., responders; n = 188) and compared with patients who failed to achieve SVR12 (i.e., non-responders; n = 109) or healthy controls (n = 62). Our results demonstrated that patients bearing HINT1 rs7728773 CT/TT (odds ratio 2.119, 95% CI 1.263-3.559, p = 0.005) and IFNL3 rs12979860 CC (odds ratio 3.995, 95% CI 2.126-7.740, p = 0.0001) were more likely to achieve SVR12. However, neither HINT1 rs4696 nor NME1 rs3760468 seems to contribute to the responsiveness to SOF/DCV. Binary regression analysis defined 5 predictor factors independently associated with SVR12: age, bilirubin, HB, early stages of fibrosis, and combined HINT1 rs7728773 and IFNL3 rs12979860 favorable and mixed genotypes (odds ratio 3.134, 95% CI 1.518-6.47, p = 0.002), and that was confirmed by the combined ROC curve for the 5 predictor factors (AUC = 0.91, 95% CI 0.869-0.95, P = 0.0001). In conclusion, these data suggest that the two SNPs have the potential in predicting the response rate to SOF/DCV treatment in patients infected with HCV GT4. This study is the first to investigate the pharmacogenetics of SOF metabolizing enzyme and introduce HINT1 rs7728773 as a novel SNP that predict the treatment efficacy.
Production of amylases by fungi under solid-state fermentation is considered the best methodology... more Production of amylases by fungi under solid-state fermentation is considered the best methodology for commercial scaling that addresses the ever-escalating needs of the worldwide enzyme market. Here response surface methodology (RSM) was used for the optimization of process variables for α-amylase enzyme production from Trichoderma virens using watermelon rinds (WMR) under solid-state fermentation (SSF). The statistical model included four variables, each detected at two levels, followed by model development with partial purification and characterization of α-amylase. The partially purified α-amylase was characterized with regard to optimum pH, temperature, kinetic constant, and substrate specificity. The results indicated that both pH and moisture content had a significant effect (P < 0.05) on α-amylase production (880 U/g) under optimized process conditions at a 3-day incubation time, moisture content of 50%, 30 °C, and pH 6.98. Statistical optimization using RSM showed R2 values of 0.9934, demonstrating the validity of the model. Five α-amylases were separated by using DEAE-Sepharose and characterized with a wide range of optimized pH values (pH 4.5-9.0), temperature optima (40-60 °C), low Km values (2.27-3.3 mg/mL), and high substrate specificity toward large substrates. In conclusion, this study presents an efficient and green approach for utilization of agro-waste for production of the valuable α-amylase enzyme using RSM under SSF. RSM was particularly beneficial for the optimization and analysis of the effective process parameters.
Xylan saccharification is a key step in many important biotechnological applications. Xylose is t... more Xylan saccharification is a key step in many important biotechnological applications. Xylose is the main product of xylan degradation and is a major xylanase inhibitor in a bioreactor; however, xylose-binding site of xylanase is not discovered yet. Evolving of xylose-tolerant xylanase variants will reduce the cost of xylanases in industry. Glycoside hydrolase family-10 thermostable Geobacillus stearothermophilus xylanase XT6 is non-competitively inhibited by xylose with inhibition constant ki equals to 12.2 mM. In the absence of X-ray crystallography of xylanase-xylose complex, unbiased random mutagenesis of the whole xylanase gene was done by error-prone polymerase chain reaction constructing a huge library. Screening a part of the library revealed xylose-tolerant mutants having three mutations, M116I, L131P and L133V, clustered in the N-terminus of α-helix 3. The best xylose-tolerant mutant showed higher ki and catalytic capability than that of the parent by 3.5- and 3-fold, respectively. In addition, kcat increased 4.5-fold and KM decreased 2-fold. The molecular docking of xylose into xylanase XT6 structure showed that xylose binds into a small pocket between N-terminus of α-helices 3 and 4 and close to the three mutations. Mobility of α-helices 3 and 4, which controls catalysis rate, is restricted by xylose binding and increased by these mutations.
Planarians have been long known for their regenerative ability, which hinges on pluripotency. Rec... more Planarians have been long known for their regenerative ability, which hinges on pluripotency. Recently, however, the planarian model has been successfully established for routine toxicological screens aimed to assess overproliferation, mutagenicity and tumorigenesis. In this study, we focused on planarian tumor suppressor genes (TSGs) and their role during chemically induced carcinogenic stress in Combining and proteomic screens with exposure to human carcinogen type 1A agent cadmium (Cd), we showed that many TSGs have a function in stem cells and that, in general, exposure to Cd accelerated the onset and increased the severity of the observed phenotype. This suggested that the interaction between environmental and genetic factors plays an important role in tumor development in Therefore, we further focused on the synergistic effects of Cd exposure and knockdown (KD) at the cellular and molecular levels. Cd also produced a specific proteomic landscape in homeostatic animals, with 17...
Heat-shock protein 110 (HSP110) family members act as nucleotide exchange factors (NEF) of mammal... more Heat-shock protein 110 (HSP110) family members act as nucleotide exchange factors (NEF) of mammalian and yeast HSP70 chaperones during the ATP hydrolysis cycle. In this study, we describe the expression pattern of murine HSPA4, a member of the HSP110 family, during testis development and the consequence of HSPA4 deficiency on male fertility. HSPA4 is ubiquitously expressed in all the examined tissues. During prenatal and postnatal development of gonad, HSPA4 is expressed in both somatic and germ cells; however, expression was much higher in germ cells of prenatal gonads. Analyses ofHspa4-deficient mice revealed that all homozygous mice on the hybrid C57BL/6J×129/Sv genetic background were apparently healthy. Although HSPA4 is expressed as early as E13.5 in male gonad, a lack of histological differences betweenHspa4−/−and control littermates suggests thatHspa4deficiency does not impair the gonocytes or their development to spermatogonia. Remarkably, an increased number of theHspa4-de...
Journal of Molecular and Cellular Cardiology, 2012
Failure of molecular chaperones to direct the correct folding of newly synthesized proteins leads... more Failure of molecular chaperones to direct the correct folding of newly synthesized proteins leads to the accumulation of misfolded proteins in cells. HSPA4 is a member of the heat shock protein 110 family (HSP110) that acts as a nucleotide exchange factor of HSP70 chaperones. We found that the expression of HSPA4 is upregulated in murine hearts subjected to pressure overload and in failing human hearts. To investigate the cardiac function of HSPA4, Hspa4 knockout (KO) mice were generated and exhibited cardiac hypertrophy and fibrosis. Hspa4 KO hearts were characterized by a significant increase in heart weight/body weight ratio, elevated expression of hypertrophic and fibrotic gene markers, and concentric hypertrophy with preserved contractile function. In response to pressure overload, cardiac hypertrophy and remodeling were further aggravated in the Hspa4 KO compared to wild type (WT) mice. Cardiac hypertrophy in Hspa4 KO hearts was associated with enhanced activation of gp130-STAT3, CaMKII, and calcineurin-NFAT signaling. Protein blot and immunofluorescent analyses showed a significant accumulation of polyubiquitinated proteins in cardiac cells of Hspa4 KO mice. These results suggest that the myocardial remodeling of Hspa4 KO mice is due to accumulation of misfolded proteins resulting from impaired chaperone activity. Further analyses revealed a significant increase in cross sectional area of cardiomyocytes, and in expression levels of hypertrophic markers in cultured neonatal Hspa4 KO cardiomyocytes suggesting that the hypertrophy of mutant mice was a result of primary defects in cardiomyocytes. Gene expression profile in hearts of 3.5-week-old mice revealed a differentially expressed gene sets related to ion channels, muscle-specific contractile proteins and stress response. Taken together, our in vivo data demonstrate that Hspa4 gene ablation results in cardiac hypertrophy and fibrosis, possibly, through its role in protein quality control mechanism.
Background Molecular chaperones assist protein folding, facilitate degradation of misfolded polyp... more Background Molecular chaperones assist protein folding, facilitate degradation of misfolded polypeptides, and thereby maintain protein homeostasis. Impaired chaperone activity leads to defective protein quality control that is implicated in multiple skeletal muscle diseases. The heat shock protein A4 (HSPA4) acts as a co-chaperone for HSP70. Previously, we showed that Hspa4 deletion causes impaired protein homeostasis in the heart. However, its functional role in skeletal muscle has not been explored. Methods We performed a comparative phenotypic and biochemical analyses of Hspa4 knockout (KO) mice with wild-type (WT) littermates. Results HSPA4 is markedly upregulated in regenerating WT muscle in vivo, and in differentiated myoblasts in vitro. Hspa4-KO mice are marked by growth retardation and increased variability in body weight, accompanied by 35% mortality rates during the peri-weaning period. The surviving Hspa4-KO mice experienced progressive skeletal muscle myopathy, character...
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Papers by Amal Zohir