• Food Science of Animal Resources
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• v.31 no.2
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• pp.241-249
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• 2011

This study was conducted to analyze ash content, mineral composition, hydroxy methyl furfural (HMF) content, stable carbon isotope ratio, and SDS-polyacrylamide gel electrophoresis patterns to investigate the quality characteristics of various honeys harvested from different sources and to identify differences useful for distinguishing honey sources. Ash content was 0.046-0.012% in acacia honey, 0.565-1.318% in chestnut honey, 0.06-0.582% in polyfloral honey, and 0.237-0.893% in native bee honey. Potassium content was high in order of chestnut honey>native bee honey>polyfloral honey>acacia honey. The Na/K ratio was 0.92-1.97 in acacia honey, 0.02-1.59 in chestnut honey, 0.02-5.30 in polyfloral honey, and 0.22-0.51 in native bee honey. The HMF content was 9.60-12.85, 10.15-25.75, 9.7-33.5, and 6.25-21.5 mg/kg in acacia, chestnut, native bee, and polyfloral honeys, respectively. HMF content was the highest in native bee honey. A 59 kDa protein band was revealed in all samples by SDS-PAGE analysis. Protein bands of 32.1, 31.9, and 33.5 kDa were revealed in some chestnut honeys, and protein bands of 32.3 and 32.5 kDa were shown in native bee honeys. A protein band of 72 kDa was also confirmed in some chestnut honeys.

• Korean Journal of Food Science and Technology
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• v.31 no.6
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• pp.1635-1641
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• 1999

Bacillus subtilis PCA20-3 was isolated from meju and was found to produce a protease. The strain produced the maximum amount of enzyme in the medium containing soytone (0.2%), soluble starch (2%), $(NH_4)_2SO_4\;(0.1%),\;CaCl_2(0.1%),\;yeast\;extract\;(0.01%),\;K_2HPO_4\;(0.1%),\;and\;KH_2PO_4\;(0.1%)$. Protease was first concentrated by ammonium sulfate (80% saturation, w/v) precipitation of culture supernatant. Then the enzyme was purified by column chromatography using CM Sephadex C-50. The collected proteins were rechromatographed using Sephadex G-100 gel filtration column. The fraction with protease active from Sephadex G-100 gel chromatography was found to be pure when examined by SDS-polyacrylamide gel electrophoresis and YMC-pak reverse phase chromatography. Specific activity, yield and purity were 76 U/mg. 2.7%, and 7.6 fold, respectively. The molecular weight of the enzyme was estimated to be 31.5 kDa by SDS-PAGE. The number of amino acids calculated from molecular weight was evaluated about 321 residues. N-terminal sequence of the enzyme was $Val^1-Pro^2-Tyr^3-Gly^4-Val^5-Ser^6-Gln^7-Gly^8-Lys^9-Ala^{10}$.

• Korean Journal of Clinical Laboratory Science
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• v.37 no.2
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• pp.78-83
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• 2005

The optimum conditions of Cordyceps militaris mycelial growth, purification and characteristics of branched-chain amino acid aminotransferase [BCAT(EC 2.6.1.42)] in this mycelium were studied. Optimum pH, temperature and medium of culture of mycelia were 5.5, $22.5^{\circ}C$ and Hamada medium (HM), respectively. BCAT in homogenate of this mycelia was precipitated by 20-40% saturated solution of ammonium sulfate and then purified by DEAE (diethylaminoethyl)-Sephadex A-50 column chromatography with linear concentration gradient and Sephadex G-200 gel filtration. A single band of purified enzyme was detected on SDS-PAGE (sodium dodecylsulfate-polyacrylamide gel electrophoresis). Optimum pH and temperature of BCAT were found to be 7.8 and $29^{\circ}C$, respectively. It showed activity toward L-leucine, L-isoleucine and L-valine as a substrate. The Km values of this enzyme for L-leucine were determined to be 5.88 mM for L-leucine.

• Microbiology and Biotechnology Letters
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• v.23 no.1
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• pp.60-67
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• 1995

Methanol assimilating yeast, Hansenula sp. MS-364 that has high productivity with methanol as carbon and energy source has been preserved at dept. of Microbiological engineering. Purification and properties of alcohol oxidase (E.C.1.1.3.13: oxygen oxidoreductase) were investigated in the methanol assimilating yeast, Hansenula sp. MS-364. Alcohol oxidase is related to the catalytic reaction that degrades alcohol to aldehyde and peroxide. The methanol oxidizing enzyme was purified by ammonium sulfate precipitation, DEAE-Sephadex A-50 chromatography and gel filtration on Sepharose 6B from cell-free extract. The purified enzyme preparation gave a single band in the sodium dodesyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of the enzyme was calculated to be about 576,000 and molecular weight of subunit was also calculated to be 72,000. The optimal pH and temperature of the enzyme reaction were pH 7.5 and 37$\circ$C, respectively. The enzyme was unstable in acidic pH and higher temperature. The enzyme was not specific for methanol and also oxidized lower primary alcohols. The Km value for methanol was 2.5 mM and that for ethanol was 1.66 mM. The enzyme was heavily inhibited by metal ions such as Hg$^{2+}$, Ag$^{2+}$, Cu$^{2+}$. The high concentration of EDTA and sulfhydryl reagents strongly inhibited the enzyme activity. The component of coenzyme was determined to flavin adenine dinucleotide.

• Journal of Microbiology and Biotechnology
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• v.20 no.8
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• pp.1215-1218
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• 2010

The recombinant DNA pLR5cat_PSAB, in which pediocin PA-1 structural and immunity genes (pedAB) fused with the promoter and deduced signal sequence of an ${\alpha}$-amylase gene from a bifidobacterial strain were inserted in Escherichia coli-lactobacilli shuttle vector pLR5cat, was transferred to Lactobacillus reuteri KCTC 3679 and the transformant presented bacteriocin activity. The recombinant L. reuteri KCTC 3679 transformed with the shortened pLR5cat(S)_PSAB, where a nonessential region for the lactobacilli replicon was removed, also showed bacteriocin activity. The molecular mass of the secreted pediocin PA-1 from the recombinant bacteria was the same as that of native pediocin PA-1 (~4.6 kDa) from Pediococcus acidilactici K10 on a sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel. In cocultures with Listeria monocytogenes, the recombinant L. reuteri KCTC 3679 effectively reduced the viable cell count of the pathogenic bacterium by a 3 log scale compared with a control where L. monocytogenes was incubated alone.

• Applied Biological Chemistry
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• v.31 no.4
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• pp.382-386
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• 1988

Extracellular protein produced by Bacillus sp. $T_2-3$ was characterized for its patterns of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and sephadex G-100 filtration, spectrum of maximum absorption, composition of amino acid and solubility to solvents. The extracellular protein was composed of two kinds of protein which was little difference in molecular weight about 49,000. Maximum absorbance of the extracellular protein was showed at 230nm and main amino acids of the extracellular protein were aspartic acid, glutamic acid and alanine. Solubility of the extracellular protein was 55.8% in $H_2O$ and 28.4% in 0.4% NaOH.

• Fisheries and Aquatic Sciences
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• v.2 no.2
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• pp.218-225
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• 1999

A protease without tryptic and chymotryptic activities was purified from the hepatopancreas of shrimp, Penaeus orientalis, using Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, Mono-Q, and gel chromatography. Molecular weight (M.W.) of the protease was estimated to be 27kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS­PAGE). The amino acid composition of the protease was different from that of protease from P. japonicus or trypsin from P. orientalis. The protease was completely inhibited by benzamidine, $N\alpha-p-tosyl-L-lysine$ chloromethyl ketone (TLCK), and phenylmethylsulfonyl fluoride (PMSF) and was not affected by leupeptin, pepstatin, N-tosyl-L-phenylalanine chloromethyl ketone (TPCK), iodoacetate, and ethylenediamine tetra acetate (EDTA). The enzyme did not have any activity against Na-benzoyl-DL-arginine p-nitroanilide (BAPNA) or N-benzoyl-L-tyrosine ethyl ester (BTEE) which are specific substrates of trypsin and chymotrypsin, respectively. However, the protease showed hydrolytic activity for a carboxyl terminal of Tyr, Trp, Phe, Glu, and Cys.

• BMB Reports
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• v.28 no.1
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• pp.6-11
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• 1995

A new lectin, named TRA, was purified from Trichosanthes kirilowii root by acid-treated Sepharose 6B, Mono-Q, and TSK-gel 3000SW column sequential chromatography. The lectin appeared homogeneous by native gel electrophoresis at pH 4.3 and gave two protein bands of Mr=31 and 28 kDa by SDS-PAGE. The N-terminal amino acid sequences of the polypeptides of TRA have not been reported in amino acid sequences of the lectins. TRA lectin formed a precipitate with asialofetuin, neuraminidase-treated fetuin. A sugar inhibition assay indicated that N-acetyl-D-galactosamine, among the monosaccharides tested, was the most potent inhibitor of TRA-induced hemagglutination. Asialofetuin showed a 260-times stronger inhibitory activity than N-acetyl-D-galactosamine. TRA lectin also showed agglutination with normal leukocytes and lymphoma cells, but not with premature hemopoietic cells. These results suggest that TRA is a novel plant lectin.

• Mycobiology
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• v.35 no.4
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• pp.219-225
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• 2007

A keratinolytic enzyme secreted by Aspergillus flavus K-03 cultured in feather meal basal medium (FMBM) containing 2% (w/v) chicken feather was purified and characterized. Keratinolytic enzyme secretion was the maximal at day 16 of the incubation period at pH 8 and $28^{\circ}C$. No relationship was detected between enzyme yield and increase of fungal biomass. The fraction obtained at 80% ammonium sulfate saturation showed 2.39-fold purification and was further purified by gel filtration in Sephadex G-100 followed by ion exchange chromatography on DEAE-Sephadex A-50, yielding an active protein peak showing 11.53-fold purification. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and zymograms indicated that the purified keratinase is a monomeric enzyme with 31 kDa molecular weight. The extracellular keratinase of A. flavus was active in a board range of pH ($7{\sim}10$) and temperature ($30^{\circ}C{\sim}70^{\circ}C$) profiles with the optimal for keratinase activity at pH 8 and $45^{\circ}C$. The keratinase activity was totally inhibited by protease inhibitors such as phenylmethylsulfonyl fluoride (PMSF), iodoacetic acid, and ethylenediaminetetraacetate (EDTA) while no reduction of activity by the addition of dithiothreitol (DTT) was observed. N-terminal amino acid sequences were up to 80% homologous with the fungal subtilisins produced by Fusarium culmorum. Therefore, on the basis of these characteristics, the keratinase of A. flavus K-03 is determined to be subtilisins-like.

• Fisheries and Aquatic Sciences
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• v.1 no.2
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• pp.209-215
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• 1998

Trypsin-like enzyme was purified from shrimp hepatopancreas through Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, and Superdex 75 gel chromatography. Purity of trypsin-like enzyme was increased 69-fold with $44\%$ yield. The enzyme consisted of a single polypeptide chain with a molecular weight (M.W.) of 32 kDa judged by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was completely inactivated by serine enzyme inhibitors such as soybean trypsin inhibitor (SBTI), tosyl-L­lysine chloromethyl ketone (TLCK), and leupeptin. However, the enzyme was not affected by tosyl-L-phenylalanine chloromethyl ketone (TPCK) which is a chymotrypsin specific inhibitor. The enzyme had no activity against benzoyl-tyrosine ethyl ester (BTEE) which is a chymotrypsin specific substrate. The enzyme showed high activity on the carboxyl terminal of Phe, Tyr. Glu, Arg, and Asp. However. no activity was detected against the carboxyl terminal of Pro, Trp, Cys, Gly, Val, and Ala.