• Korean Journal of Microbiology
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• v.32 no.2
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• pp.155-162
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• 1994

Cells of Acinetobacter sp. strain JC1 DSM 3803, an aerobic monoxide-oxidizing bacterium, growing on glucose exhibited high catalase activity at the mid-exponential growth phase. The enzyme activity decreased gradually after then until the early stationary phase, increased again at the mid-stationary phase, and then decreased again thereafter. Cells growing on glucose was found to contain three kinds of catalses. Cat1, Cat2 and Cat3. The activities of Cat1 and Cat3 did change significantly during growth, but that of Cat2 exhibited significant variation. Cat3 was found to present only in cells growing on glucose, but not in cells growing on carbon monoxide of methanol. The activities of call and Cat3 in cell-free extracts were stable upon treatment with ethanol and chloroform, but decreased to some extent when the enzymewere treated with 2mM $H_2O_2$ and/or 3-amino-1,2,4-triazole (AT). Cat2 was found to be extremely sensitive to the ethanol-chloroform and $H_2O_2$ treatments, but was insensitive to the AT treatment. Cat1 exhibited enzyme activity after incubation for 1 min at 80$^{\circ}C$. Cat2 and Cat3 did not show enzyme activity after incubation for 1 min at 60$^{\circ}C$ and 70$^{\circ}C$, respectively. Cat2 was found to have peroxidase activity. Cat3 was purified to homogenity in seven steps. The molecular weight of the native enzyme was estimated to be 150,000. Sodium dodecyl sulfate-gel electrophoresis revealed two identical subunits of molecular weight 65,000. The enzyme was found to show two $K_m$ values of 39 mM and 58mM. The optimal pH for the enzyme activity was 7.0, but the activities at pH 6.0, 8.0, and 9.0, were found to be comparable to that at the optimal pH. The optimal temperature for the enzyme activity was found to be 40$^{\circ}C$. The enzyme also exhibited strong activity at 20$^{\circ}C$, 30$^{\circ}C$, and 50$^{\circ}C$. The purified enzyme was not affected by the ethanol-chloroform treatment. The enzyme, howerver, showed less than 10% of the original activity when it was treated with 12 mN AT, 0.1 mM $NaN_3$ of 1mM KCN.

• Korean Journal of Microbiology
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• v.40 no.1
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• pp.29-36
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• 2004

The purpose of this work was to investigate the relationships between acrylamide degradation by Pseudomonas sp. JK-7 and several relevant physicochemical environment parameters. In initial experiments, the bacterial culture, strain JK-7 isolated from paddy soil sample was developed to grow aerobically with acrylamide as the sole source of carbon and nitrogen. The bacterium was identified as genus Pseudomonas in the basis of use BIOLOG test, and designated as Pseudomunas sp. JK-7. Strain JK-7 could degrade 50 mM acrylamide completely within 72 hours of incubation. Major intermediates resulting from acrylamide degradation were not detected with the HPLC methodology except acrylic acid which appeared to accumulate transiently in the growth medium. The pH increased from 7.0 to 8.7 with complete degradation of the initial 50 mM acrylamide within 72 hours of incubation. pH control in the range of 5 to 9 influenced the growth of JK-7 and acrylamide degradation, whereas it was not examined the growth and degradation at pH 3 or pH 11, respectively. The effect of supplemented carbons (e.g., glucose, fructose, citrate, succinate) on the acrylamide degradation by the test culture of JK-7 was evaluated. The results indicated that the addition of carbons accelerated the bacterial growth and acrylamide degradation compared to those in the absence of supplemented carbons. The effect of supplemented nitrogens on the degradation was monitored. Increasing concentrations of yeast extract resulted in higher growth yield, based on the turbidity measurement, and complete degradation of acrylamide. However, acrylamide degradation was essentially uninfluenced by the addition of $(NH_{4})_{2}SO_{4}$, $NH_4Cl$ or urea. Addition of $AgNO_3$, $CuSO_4$ or $HgCl_2$ except $ZnSO_4$ in the test culture inhibited the degradation of acrylamide and growth of JK-7.

• Korean Journal of Microbiology
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• v.38 no.3
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• pp.153-161
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• 2002

Sox4, a transcription factor, consists of three functional domains: an HMG-box domain as a DNA binding domain, serine rich region as a transactivation domain and glycine rich region (GRR), an unknown functional domain. Although Sox4 is known to be functionally involved in heart, B-cell and reproductive system development, its physiological function remains to be elucidated. We used pGEX expression system to develop a simple and rapid method for purifying Sox4 protein in suitable forms for biochemical studies of their functions. Unexpectedly, we observed that full-length Sox4 appears to be protease-sensitive during expression and purification in E. coli. To map the protease-sensitive site in Sox4, we generated various constructs with each of functional domains of Sox4 and purified as the GST-Sox4 fusion proteins using glutathione beads. We found that the specific cleavage site for the proteolytic enzyme, which exists in E. coli, is localized within the novel GRR of Sox4. Our study suggest that the GRR of Sox4 may a target for the cellular protease action and this cleavage in the GRR may be involved in regulating physiological function of Sox4. Additionally, our study may provide a useful method for investigating the proteolytic cleavage of the target molecule in E. coli.

• Korean Journal of Microbiology
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• v.33 no.4
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• pp.262-266
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• 1997

Total, direct viable count, and acid-tolerant epiphytic bacterial population sizes were quantified on leaves of chestnut tree (Castanea crenata S. et Z.) near Taejon Industrial Estate affected by acid precipitation and deposition as well as in the clean natural forest area, Mt. Kyejok, in Taejon city from August 1996 to August 1997. Geometric mean numbers of total, direct viable count, and acid-tolerant epiphytic bacteria were $9.9{\times}10^5cell/cm^2$, $1.6{\times}10^6cell/cm^2$, and $7.1{\times}10^3cfu/cm^2$ respectively, being 1.5, 2, and 2.6 times those in the clean area. Acid-tolerant epiphytic bacterial numbers at pH 5.6 by MPN method were $3.3{\times}10^4$ in the industrial area, about the same as the number, $3.4{\times}10^4MPN/cm^2$, of the clean area. Acid-tolerant bacterial number at pH 4.0 was $1.9{\times}10^{-1}MPN/cm^2$ in the industrial area, whereas none was detected in the clean area. Acid-tolerant bacteria at pH 3.0 were not detected at all in the industrial area as well as in the clean area. Epiphytic bacterial population sizes were generally the greatest in May when leaves are emerged and grew hut the lowest in November when defoliation occurs. These results showed that air pollutant deposition on leaves did not cause a decrease of epiphytic bacteria at least and acid deposition on leaves did cause an increase of acid-tolerant bacteria.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.356-363
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• 2011

The accidental releases of total petroleum hydrocarbons (TPH) due to oil spills frequently ended up with soil and ground water pollution. TPH may be degraded through physicochemical and biological processes in the environment but with relatively slow rates. In this study an attempt has been made to develop an integrated chemical and biological treatment technology in order to establish an efficient and environment-friendly restoration technology for the TPH contaminated soils. A Fenton-like reaction was employed as a preceding chemical treatment process and a bioaugmentation process utilizing a diesel fuel degrader consortium was subsequently applied as a biological treatment process. An efficient chemical removal of TPH from soils occurred when the surfactant OP-10S (0.05%) and oxidants ($FeSO_4$ 4%, and $H_2O_2$ 5%) were used. Bioaugmentation of the degrader consortium into the soil slurry led to an increase in their population density at least two orders of magnitude, indicating a good survival of the degradative populations in the contaminated soils ($10^8-10^9$ CFU/g slurry). TPH removal efficiencies for the Fenton-treated soils increased by at least 57% when the soils were subjected to bioaugmentation of the degradative consortium. However, relatively lower TPH treatment efficiencies (79-83%) have been observed in the soils treated with Fenton and the degraders as opposed to the control (95%) that was left with no treatment. This appeared to be due to the presence of free radicals and other oxidative products generated during the Fenton treatment which might inhibit their degradation activity. The findings in this study will contribute to development of efficient bioremediation treatment technologies for TPH-contaminated soils and sediments in the environment.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.289-294
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• 2011

We investigated the cold shock sensitivity of DEAD-box RNA helicase gene deleted strains of in Bacillus subtilis CU1065. To understand cold shock effects, cells were cultivated at $37^{\circ}C$ to log phase ($O.D_{600}$=0.5-0.6) and then temperature was shifted to $15^{\circ}C$. Cold shock slow down the growth rate of wild type and deleted strains of DEAD-box RNA helicase gene (ydbR, yfmL, yqfR, deaD). The growth rate of ydbR deleted strain is 5 times severely reduced compared to that of wild type strain (CU1065). But the growth rate of other three (yfmL, yqfR, deaD) deleted strains is nearly equal to the growth rate of wild type. Compared to $37^{\circ}C$, the amount of ydbR and yqfR mRNA transcripts are increased at the growth temperature of $15^{\circ}C$. On the other hands the mRNA transcripts of yfmL and deaD are not changed at both conditions of $37^{\circ}C$ and $15^{\circ}C$. Upon cold shock treatment ydbR mRNA transcript is clearly increased. After treatment of rifampicin (bacteria transcription inhibitor) the amount of ydbR mRNA was measured. Temperature shift from $37^{\circ}C$ to $15^{\circ}C$ and rifampicin treatment showed slowly decay of ydbR mRNA. But at $37^{\circ}C$ and rifampicin treatment ydbR mRNA is rapidly reduced. These results showed that cold shock induction of ydbR mRNA resulted from the stability of ydbR mRNA and not from the transcription induction of ydbR. In relation to these results, we found the cold box element of csp (cold shock protein gene) in 5' untranslated region of ydbR gene. Cold shock induction of ydbR is caused by the stability of ydbR mRNA like the stability of csp mRNA.

• Korean Journal of Microbiology
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• v.53 no.2
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• pp.83-96
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• 2017

A constructed sea stream in Yeongdo, Busan, Republic of Korea is mostly static due to the lifted stream bed and tidal characters, and receives domestic wastewater nearby, causing a consistent odor production and water quality degradation. Bioaugmentation of a microbial consortium was proposed as an effective and economical restoration technology to restore the polluted stream. The microbial consortium activated on site was augmented on a periodic basis (7~10 days) into the most polluted site (Site 2) which was chosen considering the pollution level and tidal movement. Physicochemical parameters of water qualities were monitored including pH, temperature, DO, ORP, SS, COD, T-N, and T-P. COD and microbial community analyses of the sediments were also performed. A significant reduction in SS, COD, T-N, and COD (sediment) at Site 2 occurred showing their removal rates 51%, 58% and 27% and 35%, respectively, in 13 months while T-P increased by 47%. In most of the test sites, population densities of sulfate reducing bacterial (SRB) groups (Desulfobacteraceae_uc_s, Desulfobacterales_uc_s, Desulfuromonadaceae_uc_s, Desulfuromonas_g1_uc, and Desulfobacter postgatei) and Anaerolinaeles was observed to generally decrease after the bioaugmentation while those of Gamma-proteobacteria (NOR5-6B_s and NOR5-6A_s), Bacteroidales_uc_s, and Flavobacteriales_uc_s appeared to generally increase. Aerobic microbial communities (Flavobacteriaceae_uc_s) were dominant in St. 4 that showed the highest level of DO and least level of COD. These microbial communities could be used as an indicator organism to monitor the restoration process. The alpha diversity indices (OTUs, Chao1, and Shannon) of microbial communities generally decreased after the augmentation. Fast uniFrac analysis of all the samples of different sites and dates showed that there was a similarity in the microbial community structures regardless of samples as the augmentation advanced in comparison with before- and early bioaugmentation event, indicating occurrence of changing of the indigenous microbial community structures. It was concluded that the bioaugmentation could improve the polluted water quality and simultaneously change the microbial community structures via their niche changes. This in situ remediation technology will contribute to an eco-friendly and economically cleaning up of polluted streams of brine water and freshwater.

• Korean Journal of Microbiology
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• v.54 no.1
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• pp.9-17
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• 2018

To investigate the role of bkdR, sigL, yplP, and des genes which were known to be involved in fatty acid synthesis and sensitive at low temperature, deletion mutants of Bacillus subtilis CU1065 and JH642 were constructed. To determine the low temperature sensitivity of these genes, we compared the growth curves of cells at $37^{\circ}C$ and $15^{\circ}C$. At $37^{\circ}C$, wild type and deletion mutants showed almost similar growth but only bkdR deletion strain at $15^{\circ}C$ showed very slow growing compared with wild type. At $15^{\circ}C$ sigL and yplP deletions were somewhat slower or similar to those of wild type strain. Double and triple mutants for bkdR, sigL, yplP deletions were constructed and grown at $20^{\circ}C$ in LB agar to investigate cold sensitive growth. Double or triple deletions including bkdR deletion showed cold sensitive growing. In order to identify more clearly cold sensitive growth, the experiments were carried out under cold shock conditions in which the temperature was lowered from $37^{\circ}C$ to $15^{\circ}C$ at the point of 0.4 optical densities at 600 nm. In these cold shock experiments, only bkdR deletion showed significantly lower growing and additional des deletion increases cold sensitivity. The bkdR activates the bkd operon, which catabolized isoleucine, valine and leucine, amino acids and produce precursors for the synthesis of branched fatty acids. At cold shock growing of bkdR deletion strain, isoleucine recovered cold sensitivity of bkdR deletion but valine did not restore cold sensitivity. Isoleucine is used as a precursor for the synthesis of anteiso-branched fatty acids. On the other hand, valine is used as a precursor for the synthesis of iso-branched fatty acids. This indicates that anteiso-branched fatty acid plays an important role at the cold shock condition.

• Korean Journal of Microbiology
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• v.54 no.1
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• pp.60-68
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• 2018

A cold-active and alkaline serine protease (Pro21717) was partially purified from the Antarctic marine bacterium Pseudoalteromonas arctica PAMC 21717. On a zymogram gel containing skim milk, Pro21717 produced two distinct clear-zones of approximately 37 kDa (low intensity) and 74 kDa (high intensity). These were found to have identical N-terminal sequences, suggesting they arose from an identical precursor and that the 37 kDa protease might homodimerize to the more active 74 kDa form of the protein. Pro21717 displayed proteolytic activity at $0-40^{\circ}C$ (optimal temperature of $40^{\circ}C$) and maintained this activity at pH 5.0-10.0 (optimal pH of 9.0). Notably, relative activities of 30% and 45% were observed at $0^{\circ}C$ and $10^{\circ}C$, respectively, in comparison to the 100% activity observed at $40^{\circ}C$, and this enzyme showed a broad substrate range against synthetic peptides with a preference for proline in the cleavage reaction. Pro21717 activity was enhanced by $Cu^{2+}$ and remained stable in the presence of detergent surfactants (linear alkylbenzene sulfonate and sodium dodecyl sulfate) and other chemical components ($Na_2SO_4$ and metal ions, such as $Ba^{2+}$, $Mg^{2+}$, $Ca^{2+}$, $Zn^{2+}$, $Fe^{2+}$, $K^+$, and $Na^{2+}$), which are often included in commercial detergent formulations. These data indicate that the psychrophilic Pro21717 has properties comparable to the well-characterized mesophilic subtilisin Carlsberg, which is commercially produced by Novozymes as the trademark Alcalase. Thus it has the potential to be used as a new additive enzyme in laundry detergents that must work well in cold tap water below $15^{\circ}C$.

• Korean Journal of Food Science and Technology
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• v.45 no.1
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• pp.104-110
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• 2013

This study assessed hazards at the harvest stage of strawberry farms which may cause risk to humans. A total of 216 samples were collected from 6 strawberry farms (soil culture farms: A, B, C; nutriculture farms: D, E, F) located in Western Gyeongnam. The collected samples were subjected for sanitary indicator bacteria (aerobic plate count, coliforms and Escherichia coli), major foodborne pathogens (E. coli O157:H7, Listeria monocytogenes, Salmonella spp., Staphylococcus aureus and Bacillus cereus), and fungi. The levels of APC and coliform in the soil culture farms were 1.0-6.9 and 0.4-4.6 log CFU/g (leaf, mL, hand or 100 $cm^2$), respectively. The samples obtained from the nutriculture farms were contaminated with the levels of 0.8-4.9, and 0.2-2.6 log CFU/g (leaf, mL, hand or 100 $cm^2$) of APC and coliform. However, E. coli was not detected in any samples. In major foodborne pathogens, S. aureus was detected at the level of ${\leq}$3.3 log CFU/hand in workers' hand samples and B. cereus was detected at the levels of 0.4-4.1 log CFU/g (hand or 100 $cm^2$) in soil, plants and workers' hygiene. L. monocytogenes, E. coli O157:H7 and Salmonella spp. were not detected. Fungi were detected at the levels of 1.0-5.2 and 0.2-4.4 log CFU/g (leaf, mL, hand or 100 $cm^2$) in soil culture and nutriculture farms, respectively.