To investigate the effect of lactic acid bacteria on the heavy metal adsorption from internal organs and blood, lactic acid bacteria were isolated from human feces. Some strains resistant to heavy metals were selected by incubation in agar media containing each of chrome and cadmium salts. Among them, a strain named KP-3 was ultimately chosen due to its higher growth rate in selective broth medium containing the heavy metals at the concentration of 0.01%. The strain was identified as Lactobacillus sp. based on its morphological, cultural and physiological characteristics. For evaluating the ability to prevent accumulation of heavy metals by selected Lactobacillus sp. strain in vivo, Sprague Dawley rats were fed with heavy metal salts (cadmium, chrome and lead) with or without cultured whole cells for 7 days. The amounts of heavy metals accumulated in liver, kidney and blood were analyzed. As a result, chrome was accumulated to kidney mostly, and lead was frequently found in liver and kidney. Experimental group (rats fed with lactic acid bacteria) showed less accumulation of heavy metal than control group (rats fed with saline solution). The inhibition rates of heavy metal accumulation were calculated to 41.8% (Cd), 33.4% (Cr) and 44.2% (Pb). Especially, feeding lactic acid bacteria strongly reduced accumulation of cadmium in blood. The results showed that feeding Lactobacillus sp. KP-3 could prevent the bioaccumulation of heavy metals in the living body.
An analytical method capable of predicting various stress-strain responses in axially loaded concrete confined with FRP (fiber reinforced polymers) composites in a rational manner is presented. Its underlying idea is that the volumetric expansion due to progressive microcracking in mechanically loaded concrete is an important measure of the extent of damage in the material microstructure, and can be utilized to estimate the load-carrying capacity of concrete by considering the corresponding accumulated damage. Following from this, an elastic modulus expressed as a function of area strain and concrete porosity, the energy-balance equation relating the dilating concrete to the confining device interactively, the varying confining pressure, and an incremental calculation algorithm are included in the solution procedure. The proposed method enables the evaluation of lateral strains consecutively according to the related mechanical model and the energy-balance equation, rather than using an empirically derived equation for Poisson's ratio or dilation rate as in other analytical methods. Several existing analytical methods that can predict the overall response were also examined and discussed, particularly focusing on the way of considering the volumetric expansion. The results predicted by the proposed and Samaan's bilinear equation models correlated with observed results with a reasonable degree, however it can be judged that the latter is not capable of predicting the response of lateral strains correctly due to incorporating the initial Poisson's ratio and the final converged dilation rate only. Further, the proposed method seems to have greater benefits in other applications by the use of the fundamental principles of mechanics.
Proceedings of the Korean Society for Applied Microbiology Conference
/
2002.10a
/
pp.18-25
/
2002
The pikromycin biosynthetic system in Streptomyces venezuleae is unique for its ability to produce two groups of antibiotics that include the 12-membered ring macrolides methymycin and neomethymycin, and the 14-membered ring macrolides narbomycin and pikromycin. The metabolic pathway also contains two post polyketide-modification enzymes, a glycosyltransferase and P450 hydroxylase that have unusually broad substrate specificities. In order to explore further the substrate flexibility of these enzymes a series of hybrid polyketide synthases were constructed and their metabolic products characterized. The plasmid-based replacement of the multifunctional protein subunits of the pikromycin PKS in S. venezuelae by the corresponding subunits from heterologous modular PKSs resulted in recombinant strains that produce both 12- and 14-membered ring macrolactones with predicted structural alterations. In all cases, novel macrolactones were produced and further modified by the DesVII glycosyltransferase and PikC hydroxylase leading to biologically active macrolide structures. These results demonstrate that hybrid PKSs in S. venezuelae can produce a multiplicity of new macrolactones that are modified further by the highly flexible DesVII glycosyltransferase and PikC hydroxylase tailoring enzymes. This work demonstrates the unique capacity of the S. venezuelae pikromycin pathway to expand the toolbox of combinatorial biosynthesis and to accelerate the creation of novel biologically active natural products. The polyketide backbone of rifamycin B is assembled through successive condensation and ${\beta}$-carbonyl processing of the extender units by the modular rifamycin PKS. The eighth module, in the RifD protein, contains nonfunctional DH domain and functional KR domain, which specify the reduction of the ${\beta}$-carbonyl group resulting in the C-21 bydroxyl of rifamycin B. A four amino acid substitution and one amino acid deletion were introduced in the putative NADPH binding motif in the proposed KR domain encoded by rifD. This strategy of mutation was based on the amino acid sequences of the corresponding motif of the KR domain of module 3 in the RifA protein, which is believed dysfunctional, so as to introduce a minimum alteration and retain the reading frame intact, yet ensure loss of function. The resulting strain produces linear polyketides, from tetraketide to octaketide, which are also produced by a rifD disrupted mutant as a consequence of premature termination of polyketide assembly. Much of the structural diversity within the polyketide superfamily of natural products is due to the ability of PKSs to vary the reduction level of every other alternate carbon atom in the backbone. Thus, the ability to introduce heterologous reductive segments such as ketoreductase (KR), dehydratase (DH), and enoylreductase (ER) into modules that naturally lack these activities would increase the power of the combinatorial biosynthetic toolbox. The dehydratase domain of module 7 of the rifamycin PKS, which is predicted to be nonfunctional in view of the sequence of the apparent active site, was replaced with its functional homolog from module 7 of rapamycin-producing polyketide synthase. The resulting mutant strain behaved like a rifC disrupted mutant, i.e., it accumulated the heptaketide intermediate and its precursors. This result points out a major difficulty we have encountered with all the Amycolatopsis mediterranei strain containing hybrid polyketide synthases: all the engineered strains prepared so far accumulate a plethora of products derived from the polyketide chain assembly intermediates as major products instead of just analogs of rifamycin B or its ansamycin precursors.
This is the third of several companion papers dealing with the derivation of material constants for ductile failure criteria under hydrostatic stress. It was observed that the ultimate engineering stresses and elongations at fracture from tensile tests for round specimens with various notch radii tended to increase and decrease, respectively, because of the stress triaxiality. The engineering stress curves from tests are compared with numerical simulation results, and it is proved that the curves from the two approaches very closely coincide. Failure strains are obtained from the equivalent plastic strain histories from numerical simulations at the time when the experimental engineering stress drops suddenly. After introducing the new concept of average stress triaxiality and accumulated average strain energy, the material constants of the Johnson-Cook failure criterion for critical energies of 100%, 50%, and 15% are presented. The experimental results obtained for EH-36 steel were in relatively good agreement with the 100% critical energy, whereas the literature states that aluminum fits with a 15% critical energy. Therefore, it is expected that a unified failure criterion for critical energy, which is available for most kinds of ductile materials, can be provided according to the used materials.
Pseudomonas sp. P20 was shown to be capable of degrading biphenyl and 4-chlorobiphenyl (4CB) to produce the corresponding benzoic acids wnich were not further degraded. But the potential of the strain for biodegradation of 4CB was shown to be excellent. The pcbA, B, C and D genes responsible for the aromatic ring-cleavage of biphenyl and 4CB degradation were cloned from the chromosomal DNA of the strain. In this study, the pebC and D genes specifying degradation of 2, 3-dihydroxybiphenyl (2, 3-DHBP) produced from biphenyl by the pebAB-encoded enzymes were cloned by using pBluescript SK(+) as a vector. From the pCK102 (9.3 kb) containing pebC and D genes, pCK1022 inserted with a EcoRI-HindIII DNA fragment (4.1 kb) carrying pebC and D and a pCK1092 inserted with EcoRI-XbaI fragment (1.95 kb) carrying pebC were constructed. The expression of pcbC and D' in E. coli CK102 and pebC in E. coli CK1092 was examined by gas chromatography and UV-vis spectrophotometry. 2.3-dihydroxybiphenyl was readily degraded to produce meta-cleavage product (MCP) by E. coli CK102 after incubation for 10 min, and then only benzoic acid(BA) was detected in the 24-h old culture. The MCP was detected in E. coli CK1022 containing pebC and 0 genes (by the resting cells assay) for up to 3 h after incubation and then diminished completely in 8 h, whereas the MCP accumulated in the E. coli CK1092 culture even after 6 h of incubation. The 2, 3-DHBP dioxygenases (product of pebC gene) produced by E. coli CK1, CK102, CK1023, and CK1092 strains were measured by native PAGE analysis to be about 250 kDa in molecular weight, which were about same as those of Pseudomonas sp. DJ-12, P. pseudoa1caligenes KF707, and P. putida OU83.
To yield high concentrations of protein expressed by genetically modified Escherichia coli, it is important that the bacterial strains are cultivated to high cell density in industrial bioprocesses. Since the expressed target protein is mostly accumulated inside the E. coli cells, the cellular product formation can be directly correlated to the bacterial biomass concentration. The typical way to determine this concentration is to sample offline. Such manual sampling, however, wastes time and is not efficient for acquiring direct feedback to control a fedbatch fermentation. An E. coli K12-derived strain was cultivated to high cell density in a pressurized stirred bioreactor on a pilot scale, by detecting biomass concentration online using a capacitance probe. This E. coli strain was grown in pure minimal medium using two carbon sources (glucose and glycerol). By applying exponential feeding profiles corresponding to a constant specific growth rate, the E. coli culture grew under carbon-limited conditions to minimize overflow metabolites. A high linearity was found between capacitance and biomass concentration, whereby up to 85 g/L dry cell weight was measured. To validate the viability of the culture, the oxygen transfer rate (OTR) was determined online, yielding maximum values of 0.69 mol/l/h and 0.98mol/l/h by using glucose and glycerol as carbon sources, respectively. Consequently, online monitoring of biomass using a capacitance probe provides direct and fast information about the viable E. coli biomass generated under aerobic fermentation conditions at elevated headspace pressures.
Kim, Tae-Su;Jung, Hyung-Moo;Kim, Sang-Yong;Zhang, Liaoyuan;Li, Jinglin;Sigdel, Sujan;Park, Ji-Hyun;Haw, Jung-Rim;Lee, Jung-Kul
Journal of Microbiology and Biotechnology
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v.25
no.7
/
pp.1093-1100
/
2015
Acetate and lactate in growth media are detrimental to the production of Thermus maltogenic amylase (ThMA), a heterologous protein, as well as to the growth of recombinant Escherichia coli. Only 50 mM of acetate or 10 mM of lactate reduced 90% of specific ThMA activity. In this study, mutant E. coli strains blocked in the ackA-pta or ackA-pta and ldh pathways were created, characterized, and assessed for their culture performace in 300 L-scale fermentation. The ackApta and ldh double-mutant strain formed significantly less lactate and acetate, and produced a concomitant increase in the excretion of pyruvate (17.8 mM) under anaerobic conditions. The ackA-pta mutant strain accumulated significant acetate but had an approximately 2-fold increase in the formation of lactate. The ackA-pta and ldh double-mutant strain had superior overall performance in large-scale culture under suboptimal conditions, giving 67% higher cell density and 66% higher ThMA activity compared with those of the control strain. The doublemutant strain also achieved a 179% improvement in volumetric ThMA production.
Plastic deformation of roadbed influences the stability and maintenance of concrete slab track. Long-term plastic deformation in a railway roadbed is generated primarily due to accumulated inelastic strains caused by repeated passing of trains. Prediction of cumulative plastic deformation is important in cost-effective maintenance of railway tracks as well as for the safe operation of trains. In this study, the vertical displacements in railway roadbeds with different thicknesses of reinforced roadbed were computed. Parameters of the power model for cumulative plastic strain were calibrated by using the data from triaxial tests and full-scale loading tests. Results of three-dimensional finite element analyses of standard roadbed sections provide us with design guidelines for the selection of the thickness of reinforced roadbed.
Saccharomyces cerevisiae KNU5377 (KNU5377) and S. cerevisiae ATCC24858 (ATCC24858) were exposed to $H_2SO_4$ as a stress, which was added at various concentrations to a YPD media. The growth of KNU5377 was reduced to approximately 60% in the YPD media containing 40 nm sulfuric acid when compared to the non-stressed condition. When their growth was monitored during an overnight culture, two strains, KNU5377 and ATCC24858, could not grow when exposed to over 50 mM of sulfuric acid. After a short exposure to this acid for 1 h, KNU5377 exhibited stronger resistance against $H_2SO_4$ than ATCC24858. The neutral trehalase activity of KNU5377 unchanged despite under various concentrations of $H_2SO_4$. In contrast, It at of ATCC24858 was much low at higher $H_2SO_4$concentrations. Trehalose, a non-reducing disaccharide, was maximally accumulated after a short exposure to 60 nm $H_2SO_4$ for KNU5377, but it was reduced under more severe stressful conditions. These results suggest that KNU5377 should modulate the trehalose concentrations under the severe stress condition of high sulfuric acid concentrations. The most highly induced protein in the KNU5377 exposed to sulfuric acid was found to be an approximately 23 kDa protein, which was revealed to be the 605 large subunit ribosomal protein, Ll3 by FASTA search results.
In order to investigate the efficacy of HYGBH on atopic dermatitis, various immune related factors were studied. The results and conclusions are as follows. Atopic dermatitis symptoms were improved in HYGBH treated group and significant decrease in dermatitis index were observed in 12 and 14 weeks. HYGBH treated group showed significant decrease in CD4+, CD3+/CD69+ immune cell ratio in PBMC by 18% and 40.6% respectively. HYGBH treated group showed significant decrease in CD3+, CD11b+/Gr-1+ immune cell ratio in dorsal skin by 44.6% and 53.1% respectively. HYGBH treated group showed significant decrease in IL-4, IFN-${\gamma}$ in spleen by 29.5%, 7.7% respectively. HYGBH treated group showed decrease in the expression of IL-5, IL-13, IL-17 and histamine by 21%, 9.6%, 14%, and 32.2% respectively. Also the group showed decrease in the expression of IgE by 6.8% respectively. HYGBH treated group showed significant decrease in the transcription of IL-5 and IL-3 mRNA in skin by 35.5% and 23.2% respectively. The results above indicated that treatment of HYGBH improved atopic dermatitis symptoms by anti-oxidant activity as well as immune modulation activity as a clinical evidence. Also, to increase the application of fermented oriental medicine, different fermentation conditions using various microbial strains should be accumulated as the clinical evidence in the future.
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