Gene cloning and molecular characterization of a thermostable chitosanase from Bacillus cereus TY24.

BACKGROUND: An important conceptual advance in health and the environment has been recognized that enzymes play a key role in the green processing industries. Of particular interest, chitosanase is beneficial for recycling the chitosan resource and producing chitosan oligosaccharides. Also, chitosan gene expression and molecular characterization will promote understanding of the biological function of bacterial chitosanase as well as explore chitosanase for utilizing chitosan resources. RESULTS: A chitosanase-producing bacterium TY24 was isolated and identified as Bacillus cereus. Moreover, the chitosanase gene was cloned and expressed in Escherichia coli. Sequence analysis reveals that the recombinant chitosanase (CHOE) belongs to the glycoside hydrolases 8 family. The purified CHOE has a molecular weight of about 48 kDa and the specific activity of 1150 U/mg. The optimal pH and temperature of CHOE were 5.5 and 65 °C, respectively. The enzyme was observed stable at the pH range of 4.5-7.5 and the temperature range of 30-65 °C. Especially, the half-life of CHOE at 65 °C was 161 min. Additionally, the activity of CHOE was remarkably enhanced in the presence of Mn2+, Cu2+, Mg2+ and K+, beside Ca2+ at 5 mM. Especially, the activity of CHOE was enhanced to more than 120% in the presence of 1% of various surfactants. CHOE exhibited the highest substrate specificity toward colloid chitosan. CONCLUSION: A bacterial chitosanase was cloned from B. cereus and successfully expressed in E. coli (BL21) DE3. The recombinant enzyme displayed good stability under acid pH and high-temperature conditions.

Production of surfactant-stable keratinase from Bacillus cereus YQ15 and its application as detergent additive.

BACKGROUND: With the growing concern for the environment, there are trends that bio-utilization of keratinous waste by keratinases could ease the heavy burden of keratinous waste from the poultry processing and leather industry. Especially surfactant-stable keratinases are beneficial for the detergent industry. Therefore, the production of keratinase by Bacillus cereus YQ15 was improved; the characterization and use of keratinase in detergent were also studied. RESULTS: A novel alkaline keratinase-producing bacterium YQ15 was isolated from feather keratin-rich soil and was identified as Bacillus cereus. Based on the improvement of medium components and culture conditions, the maximum keratinase activity (925 U/mL) was obtained after 36 h of cultivation under conditions of 35 °C and 160 rpm. Moreover, it was observed that the optimal reacting temperature and pH of the keratinase are 60 °C and 10.0, respectively; the activity was severely inhibited by PMSF and EDTA. On the contrary, the keratinase showed remarkable stability in the existence of the various surfactants, including SDS, Tween 20, Tween 60, Tween 80, and Triton X-100. Especially, 5% of Tween 20 and Tween 60 increased the activity by 100% and 60%, respectively. Furtherly, the keratinase revealed high efficiency in removing blood stains. CONCLUSION: The excellent compatibility with commercial detergents and the high washing efficiency of removing blood stains suggested its suitability for potential application as a bio-detergent additive.

[Relationships between phosphate solubilizing ability, pH, and organic acids of Penicillium oxalicum YTY and its mutants]. / è‰é ¸é’éœ‰YTYåŠå ¶çªå˜ä½“è§£ç£·èƒ½åŠ›ã€pHå’Œåˆ†æ³Œæœ‰æœºé ¸çš„å ³ç³».

The high-efficiency phosphate solubilizing mutants of Penicillium oxalicum YTY were screened by mutagenesis of ion beam combined with UV. We analyzed the changes and correlation of phosphate solubilizing ability, pH, and organic acid for YTY and its mutants, and examined the phosphate solubilizing mechanism of P. oxalicum YTY. The results showed that five high-efficiency mutants, P9-8, P9-9, P15-4, P15-6, and P15-7 were screened, and that the phosphate solubili-zing ability of mutants was increased by more than 60% compared with YTY. In the process of pho-sphorus solubilization, both phosphorus solubilizing ability and rate of mutants were higher than that of YTY, and the mutants pH was significantly lower than YTY. The type and content of organic acids secreted by the mutants showed some variations. All mutants and YTY could secrete lactic acid, acetic acid and oxalic acid, while P9-8 also produced citric acid. The pH and the phosphate solubilizing ability of YTY and its mutants had a significant negative correlation. Phosphate solubilizing ability with organic acid and pH were all significantly correlated for YTY and the mutants, except P15-4. Organic acids and low environmental pH reduced by organic acids were the probable mechanism for P. oxalicum to dissolve phosphorus. Radiation of ion beam combined with UV could change the type and content of organic acids of P. oxalicum YTY, and initiate other H+ releasing pathways to lower pH, and participate phosphorus dissolution. The study provided biological mate-rials and theoretical basis for the research and development of high-efficiency phosphate solubilizing P. oxalicum and understanding the phosphate solubilizing mechanism of P. oxalicum.

Clinical and imaging differences between neonates and children with pyriform sinus fistula: which is preferred for diagnosis, computed tomography, or barium esophagography?

PURPOSE: Pyriform sinus fistula (PSF) is very rare. It is difficult to make an accurate diagnosis preoperatively. The aim of this study is to summarize the characteristics of clinical and radiologic presentations in neonates and children with PSF and to assess which examination is more helpful for diagnosis. METHODS: A retrospective review was performed in 29 patients with PSF at a single center from January 2013 to September 2016. The clinical presentation, computed tomography (CT) scan/magnetic resonance imaging (MRI), and barium esophagogram were evaluated between neonates and children. RESULTS: This study comprised 14 neonates and 15 children. Neonatal patients presented with cervical cyst, dyspnea, and stridor. However, the most common presentations of children were recurrent neck abscess and thyroiditis. All the neonates and 12 the children underwent CT scan/MRI, which showed the characteristic air pocket in the cervical lesion of 11 (78.6%) neonates and 4 (33.3%) children. The positive rates of barium esophagography in the neonates and children were 27.3% and 86.7%, respectively, the difference of which was statistically significant. CONCLUSION: To obtain an accurate diagnosis of PSF, the differences in clinical and radiologic presentation suggest that CT scan/MRI is preferred for neonates, and barium esophagography is preferred for children. CT scans performed immediately after barium swallow improve the positive rate.

[The study of fermentation liquid's absorption spectra in elastase fermentation].

The present article studied the fermentation liquid's absorption spectra, bacteria growth period and elastase' production in elastase fermentation, and compared and analyzed the their relation. The results show that the changes in the absorption spectra were closely related with bacteria growth and elastase' production. The UV spectroscopic technique is helpful for detecting the change of the organic nitrogen base and enzyme. The study offers a new method to detect fermentation process and is basic for detecting fermentation process on line by UV spectroscopic technique.