• BMB Reports
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• v.34 no.4
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• pp.347-354
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• 2001

$\alpha$-Glucosidase of an extreme thermophile, Thermus caldophilus GK24 (TcaAG), was purified 80-fold from cells to a homogeneous state and characterized. The enzyme exhibited optimum activity at pH 6.5 and $90^{\circ}C$, and was stable from pH 6.0 to 85 and up to $90^{\circ}C$. The enzyme had a half-life of 85 minutes at $90^{\circ}C$. An analysis of the substrate specificity showed that the enzyme hydrolyzed the non-reducing terminal unit of $\alpha$-1,6-glucosidic linkages of isomaltosaccharides and panose, $\alpha$-1,3-glycosidic bond of nigerose and turanose, and $\alpha$-1,2-glycosidic bond of sucrose. The gene encoding the TcaAG was cloned, sequenced, and sequenced in E. coli. The nucleotide sequence of the gene encoded a 530 amino acid polypeptide and had a G+C content of 68.4% with a strong bias for G or C in the third position of the codons (93.6%). A sequence analysis revealed that TcaAG belonged to the $\alpha$-amylase family. We suggest that this monomeric, thermostable, and broad-acting $\alpha$-glucosidase is a departure from previously exhibited specificities. It is, therefore, a novel $\alpha$-glucosidase.

• Korean Journal of Poultry Science
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• v.22 no.4
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• pp.203-211
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• 1995

In order to investigate the effect of feeding live yeast on growth performance, nutrients utilization, tibia mineral deposit and intestinal microorganism changes, a growth assay was conducted with 360 broiler chicks. Treatments were four levels of yeast as 0, 0.025, 0.05 and 0.1% in 1.83% tricalcium phosphate and two levels of yeast as 0 and 0.05% in 1.15% tricalcium phosphate. The crude protein content of live yeast was 45%, and 97% of it was in the pure protein form, with 46.6% of essential amino acids and 53.4% of non-essential amino acids. Growth performance was tended to increase by feeding the yeast but there was no significant difference(P>.05). The protein digestibility was increased as the feeding level of yeast increased. However, digestibilities of fat, fiber, calcium and phosphorus were not affected by the yeast. Ash and calcium content of tibia were increased as the level of yeast increased. Total number of E. coli in small intestine was significantly decreased(P<.05) in chicks fed yeast. Total number of Lactobaci1lus was significantly increased by the yeast feeding. The changes of microorganism in cecum had the same trend with the changes of microorganism in small intestine.

• Food Science of Animal Resources
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• v.33 no.2
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• pp.149-154
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• 2013

Food hygiene practices must be maintained from farm to table in order to prevent contamination by microorganisms. This study was conducted to investigate consumer hygiene practices related to the refrigerator storage of meat, including a microbial analysis, monitoring of refrigerator temperatures and consumer surveys of female homeowners in the capital area of Korea. Home refrigerator temperatures were maintained above $5^{\circ}C$ in 26 (19.7%) of the 132 houses investigated. The percentage of the refrigerators with a total microbial count over $10^2\;CFU/100\;cm^2$ was 14.4%. No E. coli, Salmonella spp. or Listeria monocytogenes microbes were detected. However, Staphylococcus aureus was detected in 14 houses (10.6%). The only statistically significant difference in hygiene practices between the non-contamination group and contamination group was in the last time of refrigerator cleaning (p<0.01), as determined by the consumer survey. To improve food hygiene when using a refrigerator, raw materials must be packaged, meat should be stored only on a designated shelf, and cooked foods must be contained to prevent cross-contamination. The refrigerator should be cleaned regularly, at least once a month, and refrigerator thermometers should be monitored below $5^{\circ}C$ in order to keep food safe.

• Journal of Microbiology and Biotechnology
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• v.2 no.1
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• pp.56-65
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• 1992

The complete nucleotide sequence of the Bacillus circulans F-2 RSDA gene, coding for raw starch digesting a-amylase (RSDA), has been determined. The RSDA structure gene consists of an open reading frame of 2508 bp. Six bp upstream of the translational start codon of the RSDA is a typical gram-positive Shine-Dalgarno sequence and the RSDA encodes a preprotein of 836 amino acids with an Mr of 96, 727. The gene was expressed from its own regulatory region in E. coli and two putative consensus promoter sequences were identified upstream of a ribosome binding site and an ATG start codon. Confirmation of the nucleotide sequence was obtained and the signal peptide cleavage site was identified by comparing the predicted amino acid sequence with that derived by N-terminal analysis of the purified RSDA. The deduced N-terminal region of the RSDA conforms to the general pattern for the signal peptides of secreted prokaryotic proteins. The complete amino acid sequence was deduced and homology with other enzymes was compared. The results suggested that the Thr-Ser-rich hinge region and the non-catalytic domain are necessary for efficient adsorption onto raw substrates, and the catalytic domain (60 kDa) is necessary for the hydrolysis of substrates, as suggested in previous studies (8, 9).

• Journal of Microbiology and Biotechnology
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• v.21 no.9
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• pp.914-920
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• 2011

Vibrio cholerae utilizes mannitol through an operon of the phosphoenolpyruvate-dependent phosphotransferase (PTS) type. A gene, mtlD, encoding mannitol-1-phosphate dehydrogenase was identified within the 3.9 kb mannitol operon of V. cholerae. The mtlD gene was cloned from V. cholerae O395, and the recombinant enzyme was functionally expressed in E. coli as a $6{\times}$His-tagged protein and purified to homogeneity. The recombinant protein is a monomer with a molecular mass of 42.35 kDa. The purified recombinant MtlD reduced fructose 6-phosphate (F6P) using NADH as a cofactor with a $K_m$ of $1.54{\pm}0.1$ mM and $V_{max}$ of $320.8{\pm}7.81\;{\mu}mol$/min/mg protein. The pH and temperature optima for F6P reduction were determined to be 7.5 and $37^{\circ}C$, respectively. Using quantitative real-time PCR analysis, mtlD was found to be constitutively expressed in V. cholerae, but the expression was up-regulated when grown in the presence of mannitol. The MtlD expression levels were not significantly different between V. cholerae O1 and non-O1 strains.

• Journal of Ecology and Environment
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• v.29 no.4
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• pp.343-352
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• 2006

This study investigated the relationship between land cover and the water quality variables in the rivers, which are located in the Yamaguchi prefecture of West Japan. The study area included 12 catchments covering $5,809\;Km^2$. pH, dissolved oxygen, suspended solid, E. coli, total nitrogen and total phosphorus were considered as river water quality variables. Satellite data was applied to generate land cover map. For linking alterations in land cover (at whole catchment and buffer zone levels) and the river water quality variables, multiple regression modeling was applied. The results indicated that non-spatial attribute (%) of land cover types (at whole catchment level) consistently explained high amounts of variation in biological oxygen demand (72%), suspended solid (72%) and total nitrogen (87%). At buffer zone-scale, multiple regression models that were developed to represent the linkage between the alterations of land cover and the river water quality variables could also explain high level of total variations in suspended solid (86%) and total nitrogen (91%).

• Journal of Environmental Science International
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• v.16 no.3
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• pp.247-253
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• 2007

Polycyclic aromatic hydrocarbons (PAHs) are a major concern because of their potential mutagenic and carcinogenic risks to human beings. One of these harmful, yet commonly observed PAHs is pyrene. Pyrene is one of the 16 PAHs listed by the United States Environmental Protection Agency as priority pollutants. The purposes of this research are to develop a method of pretreatment for PAH contaminants prior to a typical biological treatment and to demonstrate the biodegradablity of these compounds. Since pyrene is non-polar, hexane was chosen as a solvent to effectively dissolve pyrene. Pyrene solutions were treated with ozone, as it has hish oxidation capacity and electrophilic character. The intermediates and byproducts of pyrene were dissolved in alkaline water at pH 11.4 and neutralized to test for $BOD_5$, COD, and toxicity. These solutions were further ozonated and assessed of biodegradability. The first-order rate constant to was found to be between $0.121day^{-1}$ and $0.081 day^{-1}$, depending on the duration of reozonation. The $BOD_5/COD$ ratio was found to 0.66. The toxicity test showed that after 10 min of reozonation time, the byproducts and intermediates of pyrene were within the lion-toxic range of ${\pm}10%$ inhibition for E-Coli bacteria.

• Journal of The Korean Society of Grassland and Forage Science
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• v.39 no.3
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• pp.132-140
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• 2019

Fermented total mixed ration (TMR) is a novel feed for ruminants in South Korea. The purpose of this study was to evaluate the effects of lactic acid bacteria (LAB) on the quality of TMR and in vitro ruminal fermentation. Strains of three LAB spp. (Lactobacillus plantarum, L. brevis, L. mucosae) were used in fermentation of TMR. Inoculations with the three LAB spp. lowered pH and increased concentrations of lactic acid, acetic acid, and total organic acid compared to non-LAB inoculated control (only addition of an equivalent amount of water) (p<0.05). Bacterial composition indicated that aerobic bacteria and LAB were higher. However, E. coli were lower in the fermented TMR than those in the control treatment (p<0.05). Among the treatments, L. brevis treatment had the highest concentration of total organic acid without fungus detection. Gas production, pH, and ammonia-nitrogen during ruminal in vitro incubation did not differ throughout incubation. However, ruminal total VFA concentration was higher (p<0.05) in the LAB spp. treatments than the control treatment at 48 hours. Overall, the use of L. brevis as an inoculant for fermentation of high moisture. TMR could inhibit fungi growth and promote lactic fermentation, and enhance digestion in the rumen.

• Journal of Microbiology
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• v.45 no.2
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• pp.158-167
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• 2007

In this study, we have cloned a novel cDNA encoding for a papain-family cysteine protease from the Uni-ZAP XR cDNA library of the polychaete, Periserrula leucophryna. This gene was expressed in Escherichia coli using the T7 promoter system, and the protease was characterized after partial purification. First, the partial DNA fragment (498 bp) was amplified from the total RNA via RT-PCR using degenerated primers derived from the conserved region of cysteine protease. The full-length cDNA of cysteine protease (PLCP) was prepared via the screening of the Uni-ZAP XR cDNA library using the $^{32}P-labeled$ partial DNA fragment. As a result, the PLCP gene was determined to consist of a 2591 bp nucleotide sequence (CDS: 173-1024 bp) which encodes for a 283-amino acid polypeptide, which is itself composed of an 59-residue signal sequence, a 6-residue propeptide, a 218-residue mature protein, and a long 3'-noncoding region encompassing 1564 bp. The predicted molecular weights of the preproprotein and the mature protein were calculated as 31.8 kDa and 25 kDa, respectively. The results of sequence analysis and alignment revealed a significant degree of sequence similarity with other eukaryotic cysteine proteases, including the conserved catalytic triad of the $Cys^{90},\;His^{226},\;and\;Asn^{250}$ residues which characterize the C1 family of papain-like cysteine protease. The nucleotide and amino acid sequences of the novel gene were deposited into the GenBank database under the accession numbers, AY390282 and AAR27011, respectively. The results of Northern blot analysis revealed the 2.5 kb size of the transcript and ubiquitous expression throughout the entirety of the body, head, gut, and skin, which suggested that the PLCP may be grouped within the cathepsin F-like proteases. The region encoding for the mature form of the protease was then subcloned into the pT7-7 expression vector following PCR amplification using the designed primers, including the initiation and termination codons. The recombinant cysteine proteases were generated in a range of 6.3 % to 12.5 % of the total cell proteins in the E. coli BL21(DE3) strain for 8 transformants. The results of SDS-PAGE and Western blot analysis indicated that a cysteine protease of approximately 25 kDa (mature form) was generated. The optimal pH and temperature of the enzyme were determined to be approximately 9.5 and $35^{\circ}C$, respectively, thereby indicating that the cysteine protease is a member of the alkaline protease group. The evaluation of substrate specificity indicated that the purified protease was more active towards Arg-X or Lys-X and did not efficiently cleave the substrates with non-polar amino acids at the P1 site. The PLCP evidenced fibrinolytic activity on the plasminogen-free fibrin plate test.

• Food Science of Animal Resources
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• v.25 no.1
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• pp.13-19
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• 2005

The radio-sensitivity of pathogens and the effect of irradiation on microbiologican safety and mutagenicity of meat products such as seasoned and cooked beef and ham were investigated. Samples were radiation-sterilized and inoculated at 10/sup 7/ cfu/g with each of the four pathogens including Salmonella typhimurium, Escherichia coli, Staphylcoccus aureus, and Listeria ivanovii. No viable cells of pathogens were observed in the sample irradiated with 3 kGy. The D/sub 10/ value of inoculated pathogens in seasoned and cooked beef and ham were 0.24∼0.48 and 0.39∼0.45, respectively. Results of Ames test performed with non-irradiated and irradiated seasoned and cooked beef and ham were both negative at the level of 625, 1,250, 2,500, 50,000, and! 10,000 ㎍ sample/plate, respectively. Results indicate that low dose (2∼3 kGy) irradiation is effective to ensure safety for seasoned and cooked beef and ham with toxicological wholesomeness.