• Journal of Mushroom
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• v.14 no.4
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• pp.162-167
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• 2016

Lactic acid bacteria (LAB) producing phenyllactic acid (PLA), which is known as antimicrobial compound, was isolated from button mushroom bed and the isolated LAB was identified to Lactobacillus casei by 16 rRNA gene sequence analysis. Cell-free supernatant (CFS) from L. casei was assessed for both the capability to produce the antimicrobial compound PLA and the antifungal activity against three fungal pathogens (Rhizoctonia solani, Botrytis cinerea, and Collectotricum aculatum). PLA concentration was investigated to be 3.23 mM in CFS when L. casei was grown in MRS broth containing 5 mM phenylpyruvic acid as precursor for 16 h. Antifungal activity demonstrated that all fungal pathogens were sensitive to 5% CFS (v/v) of L. casei with average growth inhibitions ranging from 34.58% to 65.15% (p < 0.005), in which R. solani was the most sensitive to 65.15% and followed by C. aculatum, and B. cinerea. The minimum inhibitory concentration (MIC) for commercial PLA was also investigated to show the same trend in the range of 0.35 mg mL-1 (2.11 mM) to 0.7 mg mL-1 (4.21 mM) at pH 4.0. The inhibition ability of CFS against the pathogens were not affected by the heating or protease treatment. However, pH modification in CFS to 6.5 resulted in an extreme reduction in their antifungal activity. These results may indicate that antifungal activities in CFS was caused by acidic compounds like PLA or organic acids rather than protein or peptide molecules.

• Journal of Microbiology and Biotechnology
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• v.14 no.6
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• pp.1142-1149
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• 2004

Streptococcus mutans has the capacity of inducing dental caries. Thus, to develop a novel way of preventing dental caries, a cell wall hydrolase-producing strain was isolated and its characteristics were investigated. Among 200 alkalophilic strains isolated from soil, 8 strains exhibited lytic activities against Streptococcus mutans. However, strain YU5215 with the highest cell wall hydrolase activity was selected for further study. Strain YU5215 was identified as a novel strain of Bacillus based on analyzing its 16S rDNA sequence and Bergey's Manual of Systematic Bacteriology, and thus designated as Bacillus mutanolyticus YU5215. The optimal conditions for the production of the cell wall hydrolase from Bacillus mutanolyticus YU5215 consisted of glucose ($0.8\%$), yeast extract ($1.2\%$), polypeptone ($0.5\%$), $K_{2}HPO_{4}\;(0.1\%$), $MgSO_{4}{\cdot}7H_{2}O$ ($0.02\%$), and $Na_{2}CO_{3}\;(1.0\%$) at pH 10.0. Bacillus mutanolyticus YU5215 was cultured at 30^{circ}C for 72 h to produce the cell wall hydrolase, which was then purified by acetone precipitation and CM-agarose column chromatography. The molecular weight of the lytic enzyme was determined as 22,700 Da by SDS-PAGE. When the cell wall peptidoglycan of Streptococcus mutans was digested with the lytic enzyme, no increase in the reducing sugars was observed, while the free amino acids increased, indicating that the lytic enzyme had an endopeptidase-like property. The amino terminus of the cell wall peptidoglycan digested by the lytic enzyme was determined as a glutamic acid, while the lytic site of the lytic enzyme in the Streptococcus mutans peptidoglycan was identified as the peptide linkage of L-Ala and D-Glu.

• Journal of Microbiology and Biotechnology
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• v.26 no.9
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• pp.1613-1619
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• 2016

Francisella tularensis is a highly virulent pathogen of humans and other mammals. Moreover, F. tularensis has been designated a category A biothreat agent, and there is growing interest in the development of a protective vaccine. In the present study, we determine the in vitro and in vivo immune responses of a subunit vaccine composed of recombinant peptides Tul4 and FopA from epitopes of the F. tularensis outer membrane proteins. The recombinant peptides with adjuvant CpG induced robust immunophenotypic change of dendritic cell (DC) maturation and secretion of inflammatory cytokines (IL-6, IL-12). In addition, the matured DCs enabled ex vivo proliferation of naive splenocytes in a mixed lymphocyte reaction. Lastly, we determined the in vivo immune response by assessment of antibody production in C57BL/6 mice. Total IgG levels were produced after immunization and peaked in 6 weeks, and moreover, Tul4-specific IgG was confirmed in the mice receiving peptides with or without CpG. Based on these results, we concluded that the recombinant peptides Tul4 and FopA have immunogenicity and could be a safe subunit vaccine candidate approach against F. tularensis.

• Journal of Nutrition and Health
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• v.38 no.9
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• pp.750-755
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• 2005

Hepcidin, a peptide hormone synthesized mainly by the liver, has been implicated as a key regulator of iron homeostasis. Results from studies with experimental animal models suggested that hepcidin levels are related with body iron status, but little data is available in human subjects. This study was conducted to determine the relationship between serum pro-hepcidin levels, blood indexes of anemia, and dietary iron intake in female college students. Serum pro-hepcidin concentrations were measured by enzyme-linked immunosorbent assay in eighty-two women with $22.1\pm0.2$ years old. Dietary intake data were collected by using the 24-hour recall method for 3 days. Mean concentrations of serum pro-hepcidin were 85.1 ng/ml$\pm$6.1(s.d.) with the range of 13.6-295.7 ng/ml. The median value of serum pro-hepcidin in the study subjects was 70.3 ng/ml. Serum pro-hepcidin concentrations were positively correlated with hemoglobin concentrations (r=0.273, p=0.013), and also with hematocrit (r=0.291, p=0.008). To examine whether the level of dietary iron intake affects serum pro-hepcidin levels, study subjects were divided into two groups according to the amounts of daily iron intake. Serum pro-hepcidin concentrations were $22\%$ lower in groups with low iron intake (${\leq}10.1$ mg/day), compared to high-iron intake group (>10.1 mg/day) . In conclusion, these data, as in agreement with findings in mice, suggest that hepcidin plays an important role in regulating iron metabolism in the human body.

• Asian-Australasian Journal of Animal Sciences
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• v.28 no.12
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• pp.1774-1783
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• 2015

Milk lysozyme is the ubiquitous enzyme in milk of mammals. In this study, the cDNA sequence of a new chicken-type (c-type) milk lysozyme gene (YML), was cloned from yak mammary gland tissue. A 444 bp open reading frames, which encodes 148 amino acids (16.54 kDa) with a signal peptide of 18 amino acids, was sequenced. Further analysis indicated that the nucleic acid and amino acid sequences identities between yak and cow milk lysozyme were 89.04% and 80.41%, respectively. Recombinant yak milk lysozyme (rYML) was produced by Escherichia coli BL21 and Pichia pastoris X33. The highest lysozyme activity was detected for heterologous protein rYML5 (M = 1,864.24 U/mg, SD = 25.75) which was expressed in P. pastoris with expression vector $pPICZ{\alpha}A$ and it clearly inhibited growth of Staphylococcus aureus. Result of the YML gene expression using quantitative polymerase chain reaction showed that the YML gene was up-regulated to maximum at 30 day postpartum, that is, comparatively high YML can be found in initial milk production. The phylogenetic tree indicated that the amino acid sequence was similar to cow kidney lysozyme, which implied that the YML may have diverged from a different ancestor gene such as cow mammary glands. In our study, we suggest that YML be a new c-type lysozyme expressed in yak mammary glands that plays a role as host immunity.

• Asian-Australasian Journal of Animal Sciences
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• v.11 no.6
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• pp.725-731
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• 1998

Nine crossbred female pigs fitted with the bladder catheters were used to investigate the effects of dietary protein form on the efficiency of absorbed nitrogen for nitrogen retention in growing pigs. Combinations of the main protein sources were corn-soybean meal (CSM; slow + slow absorption rate form), corn-hydrolyzed casein (CAS; slow + rapid absorption rate form) and corn-porcine plasma (CPL; slow + intermediate absorption rate form). All experimental diets were formulated to be isonitrogenous (CP 11%) and isocaloric (3.5 Mcal/kg) and synthetic amino acids were added to the diet as required to maintain an equivalent amino acid profile among diets. Fecal digestibility of nitrogen was not different among treatments (p > 0.10). Ingested nitrogen was absorbed with an apparent efficiency of 82% to 84%. Mean nitrogen retention in pigs fed the CSM diet was as high as for pigs fed the CPL diet (0.74 g N/kg $BW^{0.75}$ per d), which was higher than the N retention rate in pigs fed CAS diet (0.68 g/kg $BW^{0.75}$ per d; P < 0.05). Apparent biological values (ABV = 100 ${\times}$ N retention/absorbed nitrogen) were 63.3%, 58.0% and 61.6% for CSM, CAS, and CPL groups, respectively (p < 0.05). There was no difference in mean energy digestibility among treatments. The efficiency of absorbed lysine utilization was significantly different among treatments (p < 0.05). Pigs fed the CAS diet were inferior to counterparts on the other diets in utilizing absorbed lysine. The ratios of free (and small peptide-bound) to protein-bound amino acids in CSM diet differed considerably from the CAS diet. This may affect the efficiency of amino acids utilization for nitrogen retention if hydrolyzed and intact amino acid pools reach the blood at different times.

• Food Science and Biotechnology
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• v.16 no.4
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• pp.565-571
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• 2007

This study was carried out to investigate an optimum condition for the high angiotensin-l converting enzyme (ACE) inhibitory activity and the yield on enzyme concentration, casein concentration, and hydrolysis time. The optimum condition was performed by response surface methodology for acquirement of casein hydrolysate of milk which shows high ACE inhibitory activity, Among 8 tested enzymes, Protamex showed the highest activation degree with 77.03 unit/g from casein. Their hydrolysis degrees of flovourzyme 500MG, protamex, mixture from 1% casein were 85.5, 88.5, and 93.5%, respectively. The ranges of enzyme concentration (0.25-1.25%), casein concentration (2.5-12.5%), and hydrolysis time (20-100 min) as 3 independent variables through preliminary experiments of the yield of casein hydrolysate and ACE inhibitory activity, and it shows optimum response surface at a saddle point. It shows enzyme concentration (0.64%), casein concentration (8.38%), and hydrolysis time (55.81 min) in the yield aspect and showed the highest activity at enzyme concentration (0.86%), casein concentration (5.97%), and hydrolysis time (63.86 min) in ACE inhibitory aspect. The $R^2$ value of a fitted optimum formula on the hydrolysis yield was 0.9751 as the significant level of 1%. The $R^2$ value of a fitted optimum formula on ACE inhibitory activity is 0.8398, and the significance is recognized in the range of 5%.

• Journal of Microbiology and Biotechnology
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• v.21 no.7
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• pp.667-674
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• 2011

The gene for esterase (rEst1) was isolated from a new species of genus Rheinheimera by functional screening of E. coli cells transformed with the pSMART/HaeIII genomic library. E. coli cells harboring the esterase gene insert could grow and produce clear halo zones on tributyrin agar. The rEst1 ORF consisted of 1,029 bp, corresponding to 342 amino acid residues with a molecular mass of 37 kDa. The signal P program 3.0 revealed the presence of a signal peptide of 25 amino acids. Esterase activity, however, was associated with a homotrimeric form of molecular mass 95 kDa and not with the monomeric form. The deduced amino acid sequence showed only 54% sequence identity with the closest lipase from Cellvibrio japonicus strain Ueda 107. Conserved domain search and multiple sequence alignment revealed the presence of an esterase/ lipase conserved domain consisting of a GXSXG motif, HGGG motif (oxyanion hole) and HGF motif, typical of the class IV hormone sensitive lipase family. On the basis of the sequence comparison with known esterases/ lipases, REst1 represents a new esterase belonging to the class IV family. The purified enzyme worked optimally at $50^{\circ}C$ and pH 8, utilized pNP esters of short chain lengths, and showed best catalytic activity with p-nitrophenyl butyrate ($C_4$), indicating that it was an esterase. The enzyme was completely inhibited by PMSF and DEPC and showed moderate organotolerance.

• Journal of Microbiology and Biotechnology
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• v.31 no.7
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• pp.1035-1043
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• 2021

Although engineered Saccharomyces cerevisiae fermenting cellobiose is useful for the production of biofuels from cellulosic biomass, cellodextrin accumulation is one of the main problems reducing ethanol yield and productivity in cellobiose fermentation with S. cerevisiae expressing cellodextrin transporter (CDT) and intracellular β-glucosidase (GH1-1). In this study, we investigated the reason for the cellodextrin accumulation and how to alleviate its formation during cellobiose fermentation using engineered S. cerevisiae fermenting cellobiose. From the series of cellobiose fermentation using S. cerevisiae expressing only GH1-1 under several culture conditions, it was discovered that small amounts of GH1-1 were secreted and cellodextrin was generated through trans-glycosylation activity of the secreted GH1-1. As GH1-1 does not have a secretion signal peptide, non-conventional protein secretion might facilitate the secretion of GH1-1. In cellobiose fermentations with S. cerevisiae expressing only GH1-1, knockout of TLG2 gene involved in non-conventional protein secretion pathway significantly delayed cellodextrin formation by reducing the secretion of GH1-1 by more than 50%. However, in cellobiose fermentations with S. cerevisiae expressing both GH1-1 and CDT-1, TLG2 knockout did not show a significant effect on cellodextrin formation, although secretion of GH1-1 was reduced by more than 40%. These results suggest that the development of new intracellular β-glucosidase, not influenced by non-conventional protein secretion, is required for better cellobiose fermentation performances of engineered S. cerevisiae fermenting cellobiose.

• Korea Journal of Cosmetic Science
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• v.2 no.1
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• pp.11-19
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• 2020

The aim of this study is to investigate the effect of various pentapeptides on hair repair depending on the characteristics of comprising amino acids using crosslinking agents in hair. Total ten peptides were synthesized with two kinds of amino acids respectively, of which were previously categorized according to R group of the amino acids contributing to the characteristic of each peptide: STTSS (Ser-Thr-Thr-Ser-Ser), LIILL (Leu-Ile-Ile-Leu-Leu), CMMCC (Cys-Met-Met-Cys-Cys), DEEDD (Asp-Glu-Glu-Asp-Asp), RKKRR (Arg-Lys-Lys-Arg-Arg), TAMRA-STTSS, TAMRA-LIILL, TAMRA-CMMCC, TAMRA-DEEDD, and TAMRA-RKKRR. Pentapeptide alone, or pentapeptides with crosslinking agents such as polymeric carbodiimide (PCI) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) were treated to chemically damaged hair. Hair diameter and break strength (N = 40/case) were measured to calculate tensile strength of hair for computing hair repair ratio, and fluorescence yields (N = 20/case) were collected for hair treated with TAMRA-peptides. The tensile strength of hair treated with pentapeptides alone, or pentapeptides with cross-linking agents is consistent with the fluorescence yield from the microscope images of the cross-sectioned hair in vision and in numerical values. Pentapeptides consisting of hydrophobic amino acids (LIILL), amino acids with sulfur (CMMCC), and basic amino acids (RKKRR) increased the tensile strength in perm-damaged hair. Pentapeptides with no extra carboxyl/amine groups in R group of amino acids resulted in no significant differences in hair strength and fluorescence yield among hairs treated with alone and with crosslinkers. Pentapeptides with extra carboxyl groups or amine groups enabled further strengthening of hair due to increased bonds within the hair after carbodiimide coupling reaction. The hair repairs of pentapeptides with various amino acid sequences were studied using crosslinking. Depending on the physical characteristics of comprising amino acids, the restoration of damaged hair was observed with tensile strength of hair and fluorescence signals upon cross-sectioned hair in parallel to possibly understand the binding tendency of each pentapeptide within the hair.