A thermostable $\beta$-glycosidase gene, tfi $\beta$-gly, was cloned from the genomic library of Thermus filiformis Wai33 A1. ifi $\beta$-gly consists of 1,296 bp nucleotide sequence and encodes a polypeptide of 431 amino acids. It shares a strong amino acid sequence similarity with the $\beta$-glycosidases from other Thermus spp. belonging to the glycosyl hydrolase family 1. In the present study, the enzyme was overexpressed in Escherichia coli BL21 (DE3) using the pET21b(+) vector system. The recombinant enzyme was purified to homogeneity by heat treatment and a $Ni^{2+}$-affinity chromatography. Polyacrylamide gel electrophoresis (PAGE) showed that the recombinant Tfi $\beta$-glycosidase was a monomeric form with molecular mass of 49 kDa. The temperature and pH range for optimal activity of the purified enzyme were 80- $90^{\circ}C$ and 5.0-6.0, respectively. Ninety-three percent of the enzyme activity was remained at $70^{\circ}C$ after 12 h, and its half-life at $80^{\circ}C$ was 6 h, indicating that Tfi $\beta$-glycosidase is highly thermostable. Based on its K_m$, or $K_{cat}K_m$, ratio, Tfi $\beta$-glycosidase appeared to have higher affinity for $\beta$-D-glucoside than for $\beta$-D-galactoside, however, $K_{cat} for \beta$-D-galactoside was much higher than that for $\beta$-D-glucoside. The activity for lactose hydrolysis was proportionally increased at $70^{\circ}C$ and pH 7.0 without substrate inhibition until reaching 250 mM lactose concentration. The specific activity of Tfi TEX>$\beta$-glycosidase on 138 mM lactose at $70{^\circ}C$ and pH 7.0 was 134.9 U/mg. Consequently, this newly cloned enzyme appears to have a valuable advantage of conducting biotechnological processes at elevated temperature during milk pasteurization in the production of low-lactose milk.
Bulb onion cultivation area has been restricted in southern part of Korea to avoid blotting and bulb division. The traditional culture method is transplanting bare-rooted plantlet into the field at the end of summer and harvesting at the beginning of next summer. The hot weather and weak plantlets occasionally causes unstable supply of onions in autumn. In order to enlarge cultivation area and to reduce culture period, long nursery system using plug tray and spring transplanting was tried. Forty cultivars collected from Korea and Japan were nursed using 200-plug tray and transplanted to the field in spring. Marketable yield was not related to the seedling size but lodging time. Cultivar of 'Hamasodachi' was lodged early and resulted low marketable yield. Cultivar of 'Cheonjudaego' was not lodged and yielded high but not in accordance with storability. Generally early lodged cultivars showed low storability. In order to avoid rainy harvesting season, cultivars requires excessive long time for lodging is not recommended for spring culture. Using plug nursery and spring transplanting, we successfully produced marketable onions in 3 months. But immediate using of the harvested onion is recommended. The storability of produced onions showed different result among cultivars, storage of spring onion was not recommended.
Biological control by chitinolytic microorganisms is being evaluated as management options for soilborne diseases. Forty kilograms of chitin compost (CTC) and control compost (CC) were amended on tomato plots ($15m{\times}0.5m$) 7 d before transplanting to evaluate enzymatic activities and the control of Fusarium wilt. Samples were taken on day 1, 3, 5, and 7, the day 1 corresponded to the 66 d after transplanting, the day on which the initial wilting symptoms occurred in plants of CC treated plots. The chitinase activity in soil of CTC was always higher compared to the control. Pathogenesis related (PR) protein (chitinase, ${\beta}$-1, 3-glucanase and peroxidase) activities in tomato roots in CC increased every day and showed marked differences compared to CTC. Wilting symptoms (96 d after transplanting) were reduced by 25% in CTC compared to the control. Protection of tomato plant may be correlated with the high levels of soil enzyme activities resulting from the chitin compost.
Lymphotoxin ${\beta}$ receptor ($LT{\beta}R$), a member of the tumor necrosis factor receptor family, plays an important role in lymphoid tissue's architecture and organogenesis. In contrast, MLCK and ROCK play critical roles in the regulation of stress fiber (SF) formation in cells. To determine whether $LT{\beta}R$ stimulation in fibroblastic reticular cells (FRCs) is involved in these signaling pathways, myosin light chain kinase inhibitor-7 (ML-7) was used to inhibit them. ML7-treated FRCs completely blocked SFs and showed retraction and shrinkage processes comparable to those observed in agonistic anti-$LT{\beta}R$ antibody-treated cells. The inhibition of ROCK activity with Y27632-induced changes in actin cytoskeleton organization and cell morphology in FRCs. Actin bundles rearranged into SFs, and phospho-myosin light chain (p-MLC) co-localized in FRCs. We checked the level of Rho-guanosine diphosphate (RhoGDP)/guanosine triphosphate (GTP) exchange activity using FRC lysate. When $LT{\beta}R$ was stimulated with agonistic anti-$LT{\beta}R$ antibodies, Rho-GDP/GTP exchange activity was markedly reduced. Regarding $LT{\beta}R$ signaling with a focus on MLCK inhibition, we showed that the phosphorylation of MLCs was reduced by $LT{\beta}R$ stimulation in FRCs. Cytoskeleton components, such as tubulin, b-actin, and phospho-ezrin proteins acting as membrane-cytoskeleton linkers, were produced in de-phosphorylation, and they reduced expression in agonistic anti-$LT{\beta}R$ antibody-treated FRCs. Collectively, the results suggested that MLCK and ROCK were simultaneously responsible for SF regulation triggered by $LT{\beta}R$ signaling in FRCs.
Objectives : We tried to observe the fluorescence anisotropy and intensity of ethidium ion in the intercalating binding interaction between DNA and ethidium ions in the presence of berberine, and then tried to explain the effect of berberine on the intercalating interaction of ethidium ion with DNA. Methods : DNA(calf thymus DNA), berberine and ethidium bromide(EtBr) were purchased from Sigma-Aldrich Co. Proper amount of each compound was dissolved in 20 mM sodium phosphate buffer(pH 7.0) containing 100 mM of NaCl to prepare stock solutions. Collections of the fluorescence anisotropy and intensity data were performed on JASCO FP-8300 spectrofluorometer equipped with a polarizer and a Peltier temperature controller. The excitation of ethidium ion was done at 550 nm and the emission data were collected at 600 nm. For Stern-Volmer plot, the fluorescence data were collected at $18^{\circ}C$ and $30^{\circ}C$. Results : According to the results of this research, the weak competitive binding pattern between ethidium ion and berberine appeared in binding with DNA at low ratio of DNA to ethidium ion. But at high ratio of DNA to ethidium ion, this weak competition disappeared. Instead, berberine might bind to DNA by intercalating way. In other words, berberine could de-intercalate ethidium ion from DNA at low concentration of DNA relative to ethidium ion, but could not at high concentration of DNA relative to ethidium ion. In addition, the mechanism of fluorescence quenching of ethidium ion could also proceed differently, depending on the ratio of the amount of DNA to that of ethidium ion. Conclusions : The effect of berberine on the DNA-ethidium ion intercalating interaction could work differently, depending on the relative ratio of the amount of DNA to that of ethidium ion. This study also showed that fluorescence anisotropy analysis is very useful method to obtain detailed information for investigation of the complex binding interactions. In order to fully understand the mechanism of action of the pharmacological effect by berberine, studies on the effect of berberine on the action of proteins such as various enzymes closely related to berberine-induced medicinal effects should be continued.
da Silva Teofilo, Guilherme Ferreira;Lizana, Rony Riveros;de Souza Camargos, Rosiane;Leme, Bruno Balbino;Morillo, Freddy Alexander Horna;Silva, Raully Lucas;Fernandes, Joao Batista Kochenborger;Sakomura, Nilva Kazue
Animal Bioscience
/
v.35
no.5
/
pp.690-697
/
2022
Objective: This study aimed to evaluate the effect of the ad libitum and restricted feeding regimen on fasting heat production (FHP) and body composition. Methods: Twelve Hubbard broilers breeders were selected with the same body weight and submitted in two feeding regimes: Restricted (T1) with feed intake of 150 g/bird/d and ad libitum (T2). The birds were randomly distributed on the treatments in two runs with three replications per treatment (per run). The birds were adapted to the feed regimens for ten days. After that, they were allocated in the open-circuit chambers and kept for three days for adaptation. On the last day, oxygen consumption (VO2) and carbon dioxide production (VCO2) were measured by 30 h under fasting. The respiratory quotient (RQ) was calculated as the VCO2/VO2 ratio, and the heat production (HP) was obtained using the Brower equation (1985). The FHP was estimated throughout the plateau of HP 12 hours after the feed deprivation. The body composition was analyzed by dual-energy X-ray absorptiometry scanning at the end of each period. Data were analyzed for one-way analysis of variance using the Minitab software. Results: The daily feed intake was 30 g higher to T2 (p<0.01) than the T1. Also, the birds of the T2 had significatively (p<0.05) more oxygen consumption (+3.1 L/kg0.75/d) and CO2 production (+2.2 L/kg0.75/d). That resulted in a higher FHP 359±14 kJ/kg0.75/d for T2 than T1 296±17.23 kJ/kg0.75/d. In contrast, the RQ was not different between treatments, with an average of 0.77 for the fasting condition. In addition, protein and fat composition were not affected by the treatment, while a tendency (p<0.1) was shown to higher bone mineral content on the T1. Conclusion: The birds under ad libitum feeding had a higher maintenance energy requirement but their body composition was not affected compared to restricted feeding.
Journal of the Korean Institute of Landscape Architecture
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v.52
no.2
/
pp.96-109
/
2024
The purpose of this study is to propose a method for evaluating the similarity of Show gardens using Deep Learning models, specifically VGG-16 and ResNet50. A model for judging the similarity of show gardens based on VGG-16 and ResNet50 models was developed, and was referred to as DRG (Deep Recognition of similarity in show Garden design). An algorithm utilizing GAP and Pearson correlation coefficient was employed to construct the model, and the accuracy of similarity was analyzed by comparing the total number of similar images derived at 1st (Top1), 3rd (Top3), and 5th (Top5) ranks with the original images. The image data used for the DRG model consisted of a total of 278 works from the Le Festival International des Jardins de Chaumont-sur-Loire, 27 works from the Seoul International Garden Show, and 17 works from the Korea Garden Show. Image analysis was conducted using the DRG model for both the same group and different groups, resulting in the establishment of guidelines for assessing show garden similarity. First, overall image similarity analysis was best suited for applying data augmentation techniques based on the ResNet50 model. Second, for image analysis focusing on internal structure and outer form, it was effective to apply a certain size filter (16cm × 16cm) to generate images emphasizing form and then compare similarity using the VGG-16 model. It was suggested that an image size of 448 × 448 pixels and the original image in full color are the optimal settings. Based on these research findings, a quantitative method for assessing show gardens is proposed and it is expected to contribute to the continuous development of garden culture through interdisciplinary research moving forward.
Park, Kyung-Ho;Choi, Young-Sim;Kim, Hyun-Ae;Lee, Kwang-Gill;Yeo, Joo-Hong;Jung, Do-Hyun;Kim, Sung-Han;Cho, Yun-Hi
Journal of the Korean Society of Food Science and Nutrition
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v.36
no.5
/
pp.554-562
/
2007
Ceramide rich intercellular lipid lamellae are thought to be particularly important in maintaining the structural integrity of epidermal barrier. Ceramide is synthesized de novo by serine palmitoyltransferase (SPT) phospholipid intermediates, serine and palmitic acid persist within the stratum corneum. The ceramide which is synthesized is degraded with fatty acid and sphingosine by degradative enzyme ceramidase. The depletion of ceramide in stratum corneum was reported in the atopic dermatitis. As an effort to search for the dietary source for improving the level of ceramide in epidermis, the dietary effects of various-typed silk protein were compared. Seventy male NC/Nga mice, an animal model of atopic dermatitis, were divided into seven groups: group CA as an atopic control with control diet, group S: 1% crude sericin diet, group F: 1% crude fibroin diet, group PS : peptide pattern of sericin(Mw 5000), group PF: peptide pattern of fibroin (Mw 1500), group AS: manufactured the same as amino acid profile of sericin and group AF: manufactured the same as amino acid profile of fibroin. Ten male BALB/c mice were served as group C (control group) control diet. All mice were fed on diet and water ad libitum for 10 weeks. Dry skin condition was established in group CA as ceramide content was decreased. Despite a marked decrease of mRNA and prorein expression of SPT, enzyme do novo synthesis, ceramide content of group S was dramatically increased by inhibiting the mRNA and protein expression of degradative enzyme ceramidase. However, dietary supplementation of crude silk fibroin protein (group F) and in other groups that were supplemented with either amino acid or peptide type of sericin or fibroin did not increase the level of ceramide. Together, our data demonstrate that dietary supplementation of crude sericin is more effective at improving ceramide level in epidemis of NC/Nga mice.
This studies were conducted to evaluate efficiency of microbial inoculator for active composting of food wastes. The Microbial inoculators used in this studies were purchased from different comparise to evaluate their effectiveness for composting of food waste in Korea. The number of bacteria growing at $30^{\circ}C$ in commercial inoculator collected were below $91.0{\times}10^8\;CFU/g$ which were counted from well cured compost made by animal manure. The number of bacteria in commercial microbial inoculator, such as FL, VP, B9, CM and GE were higher than that of composted at $50^{\circ}C$ or $60^{\circ}C$ of incubation temperature. Fungi were counted in GR, VP and B9 as over $10^3CFU/g$ at $30^{\circ}C$ of incubation temperature, while fungi of all the commercial inoculator collected could not grown at $50^{\circ}C$ and $60^{\circ}C$. Actinomycetes in most of the these had higher number($10^5CFU/g$) than that of compost : however, it was not detected at $60^{\circ}C$ incubation temperature from all the samples collected. The amount of carbon dioxid production was order to VP>HU>B9>GE>CM>Control>Compost in the lab scale composting test with or without inoculation of commercial inoculators, however, but the difference in carbon dioxide production was similar among each treatments. The effect of inoculation on composting parmeter such as pH changes, temperature increasing and change of chemicals properties were a little among each treatments, with or without inoculation of commercial inoculator in active composting of food waste. Using commercial inoculator did not show any statistical difference in food waste composting process under various condition such as pH changes, temperature changes, etc.
The lymphotoxin ${\beta}$ receptor ($LT{\beta}R$), a member of the tumor necrosis factor receptor family, plays an important role in lymphoid tissue's architecture and organogenesis. We found that $LT{\beta}R$ stimulation induced changes in stress fibers (SFs) in fibroblastic reticular cells (FRCs). MLCK and ROCK play critical roles in the regulation of SF formation in cells. The present study was performed to investigate the antifibrotic effects on SF regulation of $LT{\beta}R$ signaling, with a focus on MLCK inhibition. The effect of $LT{\beta}R$ on the SF change was analyzed using immunoblot and fluorescence assays and agonistic $anti-LT{\beta}R$ antibody-treated FRCs. In addition, we checked the level of Rho-guanosine diphosphate (GDP)/guanosine triphosphate (GTP) exchange activity with FRC lysate. Phospho-ezrin proteins acting as membrane-cytoskeleton linkers completely de-phosphorylated in agonistic $anti-LT{\beta}R$ antibody-treated FRCs. The actin bundles rearranged into SFs, where phospho-myosin light chain (p-MLC) co-localized in FRCs. ML7-treated FRCs completely blocked SFs and showed retraction and shrinkage processes comparable to those observed in agonistic $anti-LT{\beta}R$ antibody-treated cells. Inhibition of ROCK activity induced changes in the actin cytoskeleton organization; however, some SFs remained in the cells, while they were completely disrupted by MLCK inhibition with ML7. We showed that the phosphorylation of MLC was completely abolished with $LT{\beta}R$ stimulation in FRCs. When $LT{\beta}R$ was stimulated with the agonistic $anti-LT{\beta}R$ antibody, the Rho-GDP/GTP exchange activity was reduced, however, the activity was not completely abolished. Collectively, the results illustrated that MLCK was potently responsible for the SF regulation triggered via $LT{\beta}R$ signaling in FRCs.
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