A Thermostable Aluminum-tolerant Protease Produced by FeatherDegrading Bacillus thuringiensis Isolated from Tea Plantation

2020 ◽  
Vol 27 ◽  
Author(s):  
Tianwen Wang ◽  
Chen Liang ◽  
Sha Xiao ◽  
Li Li ◽  
Hongju Xu ◽  
...  
Keyword(s):  
Circular Dichroism ◽  
Serine Protease ◽  
Organic Solvents ◽  
Biochemical Properties ◽  
Size Exclusion ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Tea Plantation ◽  
Circular Dichroïsm ◽  
Optimal Ph

Background: Proteases with keratinolytic activity are widely used in biotechnologies. The feather-degrading Bacillus thuringensis isolated from soil sample of a tea plantation produced high level of extracellular keratinase. Objective: This study aimed to analyze the properties by biochemical and enzymological methods to gain information for better utilization of the enzyme. Methods: The enzyme was purified with ion exchange and size exclusion chromatography. The substrate preference, optimal pH and temperature, and the effects of organic solvents and ions were checked. Circular dichroism was performed to compare the secondary structures of the native and apo-enzyme. Results: The enzyme worked best at 50 o C, and it was an acidic serine protease with an optimal pH of 6.2. Ions Ca2+ and Mg2+ were essential for its activity. Organic solvents and other metal ions generally deactivated the enzyme in a concentration-dependent manner. However, Mn2+ and DMSO, which were frequently reported as inhibitors of protease, could activate the enzyme at low concentration (0.01 to 2 mmol/L of Mn2+; DMSO <2%, v/v). The enzyme exhibited high resistance to Al3+, which might be explained by the soil properties of its host’s residence. Circular dichroism confirmed the contribution of ions to the structure and activity. Conclusion: The enzyme was a thermostable aluminum-tolerant serine protease with unique biochemical properties.

Evaluation of the effects of hydrophilic organic solvents on CYP3A-mediated drug-drug interaction in vitro

2006 ◽  
Vol 25 (12) ◽  
pp. 715-721 ◽  
Author(s):  
M Iwase ◽  
N Kurata ◽  
R Ehana ◽  
Y Nishimura ◽  
T Masamoto ◽  
...  
Keyword(s):  
Organic Solvent ◽  
Organic Solvents ◽  
Human Liver ◽  
Liver Microsomes ◽  
Dependent Manner ◽  
Cyp3a4 Activity ◽  
Modulation Effect ◽  
Concentration Dependent Manner ◽  
Hydroxylation Activity

This study evaluated the effects of the commonly used hydrophilic organic solvents, acetonitrile, methanol, ethanol, 1-propanol, dimethyl sulfoxide (DMSO), N,N-dimethylformamide, polyethylene glycol and propylene glycol, on CYP3A in pooled human liver microsomes, using testosterone and midazolam as substrates. Furthermore, we examined the modulation effect of organic solvents on CYP3A inhibition by ketoconazole. Testosterone 6b-hydroxylation activity was potently inhibited in the presence of DMSO and 1-propanol in a concentration-dependent manner. Midazolam 1'-hydroxylation activity, however, was weakly inhibited only by 1% of DMSO, the highest concentration used in this study. Moreover, the potency of ketoconazole to inhibit CYP3A activities was variable, depending on the organic solvent used as a dissolving solvent for ketoconazole. Our data indicate that each organic solvent had an effect on CYP3A4 activity, evaluated by both substrates with different magnitudes. Furthermore, it was shown that the effects of organic solvents on CYP3A activity are substrate-dependent. The present study also shows that methanol had little effect on either substrate.

scholarly journals Characterization of Genipin-Modified Dentin Collagen

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Hiroko Nagaoka ◽  
Hideaki Nagaoka ◽  
Ricardo Walter ◽  
Lee W. Boushell ◽  
Patricia A. Miguez ◽  
...  
Keyword(s):  
Amino Acid ◽  
Lysyl Oxidase ◽  
Biomechanical Properties ◽  
Biochemical Properties ◽  
Cross Linking ◽  
Dependent Manner ◽  
Cross Link ◽  
Concentration Dependent Manner ◽  
Cross Links ◽  
Dentin Collagen

Application of biomodification techniques to dentin can improve its biochemical and biomechanical properties. Several collagen cross-linking agents have been reported to strengthen the mechanical properties of dentin. However, the characteristics of collagen that has undergone agent-induced biomodification are not well understood. The objective of this study was to analyze the effects of a natural cross-linking agent, genipin (GE), on dentin discoloration, collagen stability, and changes in amino acid composition and lysyl oxidase mediated natural collagen cross-links. Dentin collagen obtained from extracted bovine teeth was treated with three different concentrations of GE (0.01%, 0.1%, and 0.5%) for several treatment times (0–24 h). Changes in biochemical properties of NaB3H4-reduced collagen were characterized by amino acid and cross-link analyses. The treatment of dentin collagen with GE resulted in a concentration- and time-dependent pigmentation and stability against bacterial collagenase. The lysyl oxidase-mediated trivalent mature cross-link, pyridinoline, showed no difference among all groups while the major divalent immature cross-link, dehydro-dihydroxylysinonorleucine/its ketoamine in collagen treated with 0.5% GE for 24 h, significantly decreased compared to control (P< 0.05). The newly formed GE-induced cross-links most likely involve lysine and hydroxylysine residues of collagen in a concentration-dependent manner. Some of these cross-links appear to be reducible and stabilized with NaB3H4.

Toxicity of Industrially Relevant Chlorinated Organic Solvents In Vitro

2013 ◽  
Vol 32 (2) ◽  
pp. 136-145 ◽  
Author(s):  
Catherine McDermott ◽  
James J.A. Heffron
Keyword(s):  
Cell Proliferation ◽  
Organic Solvents ◽  
Caspase 3 ◽  
Free Calcium ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Chlorinated Organic ◽  
Free Calcium Concentration ◽  
Ros Formation

The cytotoxic effects of 4 industrially important chlorinated organic solvents, dichloromethane (DCM), 1,2-dichloroethane (DCE), trichloroethylene (TCE), and tetrachloroethylene (PERC) in vitro, were investigated. Jurkat T cells were exposed to the solvents individually for 72 hours and changes in reactive oxygen species (ROS) formation, cell proliferation, intracellular free calcium concentration ([Ca2+]), and caspase-3 activity were measured. There was a concentration-dependent increase in the ROS formation and intracellular free [Ca2+] following exposure to each of the solvents. This was accompanied by a decrease in the cell proliferation. Solvent potency decreased in the following order: PERC > TCE > DCM > DCE. Caspase-3 activity was increased in a concentration-dependent manner by TCE and PERC but was not significantly altered by DCM or DCE. n-Acetyl-l-cysteine pretreatment showed that changes in the intracellular free [Ca2+] and caspase-3 activity were independent of ROS formation. However, increased ROS formation did play a causal role in the decreased cell proliferation observed.

scholarly journals Fish-Derived Antifreeze Proteins and Antifreeze Glycoprotein Exhibit a Different Ice-Binding Property with Increasing Concentration

Biomolecules ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 423 ◽  
Author(s):  
Sakae Tsuda ◽  
Akari Yamauchi ◽  
N. M.-Mofiz Uddin Khan ◽  
Tatsuya Arai ◽  
Sheikh Mahatabuddin ◽  
...  
Keyword(s):  
Concentration Dependence ◽  
Thermal Hysteresis ◽  
Antifreeze Proteins ◽  
Biochemical Properties ◽  
Binding Property ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Ice Binding ◽  
Binding Function ◽  
The Difference

The concentration of a protein is highly related to its biochemical properties, and is a key determinant for its biotechnological applications. Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) are structurally diverse macromolecules that are capable of binding to embryonic ice crystals below 0 °C, making them useful as protectants of ice-block formation. In this study, we examined the maximal solubility of native AFP I–III and AFGP with distilled water, and evaluated concentration dependence of their ice-binding property. Approximately 400 mg/mL (AFP I), 200 mg/mL (AFP II), 100 mg/mL (AFP III), and >1800 mg/mL (AFGP) of the maximal solubility were estimated, and among them AFGP’s solubility is much higher compared with that of ordinary proteins, such as serum albumin (~500 mg/mL). The samples also exhibited unexpectedly high thermal hysteresis values (2–3 °C) at 50–200 mg/mL. Furthermore, the analysis of fluorescence-based ice plane affinity showed that AFP II binds to multiple ice planes in a concentration-dependent manner, for which an oligomerization mechanism was hypothesized. The difference of concentration dependence between AFPs and AFGPs may provide a new clue to help us understand the ice-binding function of these proteins.

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