• Journal of Microbiology and Biotechnology
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• v.29 no.2
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• pp.244-255
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• 2019

Xylose isomerase (XI; E.C. 5.3.1.5) catalyzes the isomerization of xylose to xylulose, which can be used to produce bioethanol through fermentation. Therefore, XI has recently gained attention as a key catalyst in the bioenergy industry. Here, we identified, purified, and characterized a XI (PbXI) from the psychrophilic soil microorganism, Paenibacillus sp. R4. Surprisingly, activity assay results showed that PbXI is not a cold-active enzyme, but displays optimal activity at $60^{\circ}C$. We solved the crystal structure of PbXI at $1.94-{\AA}$ resolution to investigate the origin of its thermostability. The PbXI structure shows a $({\beta}/{\alpha})_8$-barrel fold with tight tetrameric interactions and it has three divalent metal ions (CaI, CaII, and CaIII). Two metal ions (CaI and CaII) located in the active site are known to be involved in the enzymatic reaction. The third metal ion (CaIII), located near the ${\beta}4-{\alpha}6$ loop region, was newly identified and is thought to be important for the stability of PbXI. Compared with previously determined thermostable and mesophilic XI structures, the ${\beta}1-{\alpha}2$ loop structures near the substrate binding pocket of PbXI were remarkably different. Site-directed mutagenesis studies suggested that the flexible ${\beta}1-{\alpha}2$ loop region is essential for PbXI activity. Our findings provide valuable insights that can be applied in protein engineering to generate low-temperature purpose-specific XI enzymes.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.7
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• pp.1007-1014
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• 2019

Objective: This study was conducted to evaluate the fermentation characteristics under low mesophilic temperature of spent instant coffee ground (SICG) and to estimate the effect of fermented SICG (FSICG) as alternative feed ingredient on milk productivity of dairy cows. Methods: In the fermentation trial, fermentation of SICG was performed to investigate changes in characteristics using the microbial mixture (Lactobacillus plantarum, Saccharomyces cerevisiae, and Bacillus subtilis = 1:1:1) for 21 days at $20^{\circ}C$ under anaerobic conditions. Molasses was added at 5% of dry mass. In the animal trial, eighteen Holstein Friesian cows were used to evaluate the nutritive value of the FSICG which was fermented for 14 days under the same condition as the fermentation trial. Results: In the fermentation trial, the dry matter (DM) and organic matter content linearly decreased with fermentation time (p<0.001 and p = 0.008, respectively). The acid detergent insoluble nitrogen content linearly decreased with fermentation time (p = 0.037). The microorganism counts linearly increased for Lactobacillus plantarum, Saccharomyces cerevisiae, and Bacillus subtilis across fermentation time (p<0.001). In the animal trial, the DM intake of the control and FSICG treatment were not significantly different, as were milk yield, 4% fat corrected milk, fat-protein corrected milk, and feed to milk conversion content. Fat, protein, lactose, non-fat solids, milk urea nitrogen, and somatic cell counts were also not significantly different in milk composition between treatments. Conclusion: FSICG should be considered a sufficient substitute for cottonseed as a feed component, and 5% DM of a dietary FSICG level was appropriate for dairy cow diets.

• Journal of Korean Society on Water Environment
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• v.39 no.4
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• pp.302-315
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• 2023

Distribution characteristics and altitudinal gradient of four species (E. strigata, E. separigata, E. orientalis-sachalinensis group) of the mayfly genus Ephemera (Order Ephemeroptera) were analyzed with probability distribution models (exponential, normal, lognormal, logistic, Weibull, gamma, beta, Gumbel). Data was collected from 23,846 sampling units of 6,787 sites in Korea from 2010 to 2021. The beta distribution model showed the best fit for positively skewed E. orientalis-sachalinensis and little-skewed E. strigata along with altitudinal gradient. The reversed lognormal distribution model showed the best-fit for negatively skewed E. separigata. E. orientalis-sachalinensis distributed at the range of altitude 1~700 m (mean 251 m, median 226 m, mode 124 m, and standard deviation 161 m), E. strigata distributed at the range of altitude 5~871 m (mean 474 m, median 478 m, mode 492 m, and standard deviation 200 m), E. separigata distributed at the range of altitude 7~846 m (mean 620 m, median 659 m, mode 760 m, and standard deviation 181 m). Altitudinal habitat suitability ranges were estimated to be 42~257 m for E. orientalis-sachalinensis, 335~644 m for E. strigata, and 641~824 m for E. separigata. Based on the altitudinal spectrum of suitability and altitude-related temperature analysis results, E. orientalis-sachalinensis was estimated to be thermophilic, E. strigata to be mesophilic, and E. separigata to be thermophobic. This is the first national-scale evaluation of the altitudinal distribution of Ephemera in Korea. These results will be used in a further research study on altitudinal shift of the species of Ephemera under climate change.

• Proceedings of the Microbiological Society of Korea Conference
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• 2001.11a
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• pp.39-45
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• 2001

Extremophiles are unique microorganisms that are adapted to survive in ecological niches such as high or low temperatures, extremes of pH, high salt concentrations and high pressure. These unusual microorganisms have unique biochemical features which can be exploited for use in the biotechnological industries. Due to the high biodiversity of extremophilic archaea and bacteria and their existence in various biotopes a variety of biocatalysts with different physicochemical properties have been discovered. The extreme molecular stability of their enzymes, membranes and the synthesis of unique organic compounds and polymers make extremophiles interesting candidates for basic and applied research. Some of the enzymes from extremophiles, especially hyperthermophilic marine microorganisms (growth above $85^{\circ}C$), have already been purified in our laboratory. These include the enzyme systems from Pyrococcus, Pyrodictium, Thermococcus and Thermotoga sp. that are involved in polysacharide modification and protein bioconversion. Only recently, the genome of the thermoalkaliphilic strain. Anaerobranca gottschalkii has been completely sequenced providing a unique resource of novel biocatalysts that are active at high temperature and pH. The gene encoding the branching enzyme from this organism was cloned and expressed in a mesophilic host and finally characterized. A novel glucoamylase was purified from an aerobic archaeon which shows optimal activity at $90^{\circ}C$ and pH 2.0. This thermoacidophilic archaeon Picrophilus oshimae grows optimally at pH 0.7 and $60^{\circ}C$. Furthermore, we were able to detect thermoactive proteases from two anaerobic isolates which are able to hydrolyze feather keratin completely at $80^{\circ}C$ forming amino acids and peptides. In addition, new marine psychrophilic isolates will be presented that are able to secrete enzymes such as lipases, proteases and amylases possessing high activity below the freezing point of water.

• Korean Journal of Microbiology
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• v.49 no.1
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• pp.78-82
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• 2013

A novel Chryseobacterium sp. JK1 strain isolated from soil had been reported that this isolate produced large amount of extracellular protease at mesophilic temperature in previous study. The optimal temperature and pH of extracellular protease were $40^{\circ}C$ and 7.0, respectively, showing narrow range of optimal temperature and relatively broad activity from pH 6.0 to 9.0. In addition, the protease showed greatest activity against skim milk and lowest against bovine serum albumin (BSA). The protease strongly inhibited by ethylenediaminetetraacetic acid (EDTA), ethylene glycol tetraacetic acid (EGTA) or phenylmethylsulfonyl fluoride (PMSF), and addition of cation $Ag^+$ or $Cu^{2+}$, and slightly inhibited by $Al^{3+}$. No significant inhibition was found with pepstatin, and addition of cation, $K^+$, $Ca^{2+}$, $Na^+$, $Fe^{2+}$ or $Mg^{2+}$. On the contrary, protease was enhanced by addition of divalent cation $Mn^{2+}$ (5 mM). Zymography analysis of concentrated culture supernatant revealed two major bands at 67 and 145 kDa. These results suggest that Chryseobacterium sp. JK1 strain produced extracellular neutral serine proteases which could apply in food industry.

• Journal of Food Hygiene and Safety
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• v.4 no.2
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• pp.103-108
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• 1989

Three mushroom cannerries were selected by size which are representative vegetable processing firms in korea for monitoring microbial contamination of processing water, washing water, mushroom before and after washing through first and second washing tanks and, blanched and prolonged mushroom for certain time at room temperature. Total contamination degree was expressed as colony forming unit (CFU) of mesophilic aerobes. The contamination degree of processing water was $10^{2}\;CFU/100\;ml$ and washing water in first and second washing tank were 10 to 100 times higher than processing water. When 2.3 tons of washing water was used for washing 1 ton of mushroom, washing effect was showed by reduction of microbial load but cutting it to 1.8 tonsIl ton of mushroom, microbial load was higher than that of raw mushroom level. Blanching reduced microbial load to 50-500 CFU/g of blanched mushroom and it was not seen much increase of CFU in blanched mushroom left at room temperature for 3 hours in $16^{\circ}C$ processing water. Just after injection of $80^{\circ}C$ brine in container, CFU/ml of brine in container was $84{\times}10^{4}$ but it was increased rapidly to $20{\times}10^{7}$ after 2 hours at ambient temperature.

• Journal of Mushroom
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• v.18 no.3
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• pp.274-279
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• 2020

This study was conducted to investigate the characteristics of the medium used on the composting step, comparing the excavator agitator with the self-propelled turner. The temperature of the outdoor composting medium tended to increase rapidly after flipping in the turner. The late composting medium temperature was maintained at the excavator treatment area (farm practice), and the late composting effect progressed. During the field composting stage, various microorganisms such as Bacillus spp., Actinomycetes, fluorescent Pseudomonas spp., and filamentous fungi were distributed in the medium, and the density of aerobic bacteria involved in the decomposition of the medium was increased. Under high-temperature composting conditions, blue fungi, and mesophilic actinomycetes were inhibited or killed. Thermophilic actinomycetes, which play an important role in decomposing organic matter, showed higher densities than those observed in farm practices in the self-propelled turner process. The length of rice straw was slightly shorter when the self-propelled turner was used, and the water content did not show any significant difference between treatments. The a and b values tended to increase as the inverter was turned over. The CN ratio of the composting broth was lowered from 23.1 to 16.2 for the 5th turnover in the context of farming practices, and from 23.3 to 16.9 in the context of the self-propelled turner. The yield of each treatment was increased by 20% in 1 period, 28% in 2 periods, and 26% in 3 periods; the overall yield was 23%.

• Journal of the Korea Organic Resources Recycling Association
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• v.11 no.3
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• pp.75-86
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• 2003

In the near future, the capacity of conventional anaerobic digester is thought to be insufficient because of the increase of the total solids from expansion of intercepting sewer, sewage quantity and direct input of night soil from near apartment districts. The objectives of this study was to investigate the improvement of digestion efficiency using microbial agent(Bio-dh). The system was a pilot-scale, two-staged, anaerobic sludge digestion system. The first-stage digester was heated and mixed. The agitation velocity of the first-stage digester was 120rpm. The second-stage digester was neither heated nor mixed. The Digestion temperature was kept at $35{\pm}1^{\circ}C$ The detention time of digester was 19 days. The dosage of sewage sludge and microbial agent were $0.65m^3/day$ and $0.5{\ell}/day$, respectively. The experiments was run for 25days. Three times a week, $COD_{Mn}$ and SS of effluent, TS, VS, and biogas production rate were measured. Temperature, pH, and alkalinity were measured daily. The results were as follows ; Without microbial agent, digestion efficiencies ranged 46.0%~50.9%(mean=48.6%), with microbial agent(Bio-dh), digestion efficiencies ranged 52.8%~57.3%(mean=54.2%). Consequently, microbial agent(Bio-dh) increased the sludge digestion efficiency about 12%. Also, Without microbial agent, the mean concentration of $COD_{Mn}$ and SS of second-stage digester effluent were 1,639mg/L, 4,888mg/L respectively. With microbial agent, the mean concentration of $COD_{Mn}$ and SS of second-stage digester effluent were 859mg/L, 2,405mg/L respectively. Consequently, microbial agent(Bio-dh) increased the removal efficiency of $COD_{Mn}$ and SS about 47.6% and 50.8%, respectively.

• Microbiology and Biotechnology Letters
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• v.25 no.4
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• pp.386-390
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• 1997

The characteristics of alcohol dehydrogenase (ADH, EC 1.1.1.1, alcohol:NAD oxidoreductase) of thermotolerant alcohol-producing yeasts, Saccharomyces cerevisiae RA-74-2 and Kluyveromyces marxianus RA-912, were compared with that of mesophilic S. cerevisiae D, an industrial strain. Under anaerobic culture condition, both S. cerevisiae RA-74-2 and D had similar level of ADH activity at 30$\circ$C, and the activity of S. cerevisiae RA-74-2 at 37$\circ$C was the same level at 30$\circ$C. However, the level of ADH activity of S. cerevisiae D at 37$\circ$C decreased about 70% of that at 30$\circ$C. The level of enzyme activity of K. marxianus RA-912, which showed lower alcohol productivity than S. cerevisiae RA-74-2 and D, was about 43% of those strains at 30$\circ$C, and decreased somewhat at 37$\circ$C. The results showed a good correlation between the alcohol productivities and the level of ADH activities of these strains grown at 30$\circ$C and 37$\circ$C. And the higher heat stability of ADH of S. cerevisiae RA-74-2 than that of S. cerevisiae D seemed to reflect the ability of high temperature fermentation. Despite of its fermentation ability even at 45$\circ$C, however, the ADH of K. marxianus RA-912 showed lower heat stability than that of S. cerevisiae D. Both S. cerevisiae RA-74-2 and D showed similar patterns of two bands of ADH isozyme, and the low band of S. cerevisiae RA-74-2 moved slightly faster than that of S. cerevisiae D. The staining intensity of the bands of S. cerevisiae D at 37$\circ$C was weaker than those at 30$\circ$C. However, S. cerevisiae RA-74-2 showed no differences in total intensity of the bands of 30$\circ$C and 37$\circ$C. As the patterns of cellular proteins and ADH isozyme of K. marxianus RA-912 were different from S. cerevisiae RA-74-2 and D, K. marxianus might have its own characteristic ADH system.

• Korean journal of food and cookery science
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• v.26 no.2
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• pp.165-170
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• 2010

Although the demand of ready-to-eat (RTE) foods such as Kimbab is growing, large quantities and wide distribution of these foods is difficult due to their short shelf-life, exposed packaging with hygienic risk, and decreased quality at refrigerator temperatures. This study was undertaken to develop preservation and storage methods to extend the shelf-life of RTE rice products using microwave and packaging methods such as vacuum and modified atmosphere packages. RTE rice ball samples inoculated with Escherichia coli, Salmonella Typhimurium, Listeria monocytogenes, Staphylococcus aureus or Bacillus cereus were microwave treated for 0, 30, 60, 90 and 120 seconds. Populations of pathogens on the rice balls were significantly reduced with an increase in treatment time. There were more than 5 log reductions of all pathogens when the samples were microwave treated for 60 seconds. RTE rice balls inoculated with two pathogens (S. aureus and B. cereus) were packaged via air, vacuum, $N_2$ gas, and $CO_2$ gas following microwave treatment for 90 seconds. The initial S. aureus and B. cereus concentration before treatment was 7.60 and 6.59 log CFU/g, and these levels were reduced by 3.37 and 2.18 log CFU/g after microwave treatment. The levels of pathogens were significantly increased during storage time at room temperature. $CO_2$ packaging was the most effective at inhibiting microbial growth among the tested packaging methods. The levels of total mesophilic count, S. aureus and B. cereus after 5 days of storage were 7.7, 8.8 and 9.3 log CFU/g in air packaged samples and 2.4, 3.2 and 8.3 log CFU/g in $CO_2$ gas packaged samples, respectively. However, after 3 days of storage higher levels of B. cereus were observed in all samples, indicating that the samples were not safe to be consumed. Base on these results, microwave treatment and MAP packaging methods using $CO_2$ gas could be used as a potential method for extending the shelf-life of RTE foods.