• Journal of Life Science
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• v.16 no.7 s.80
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• pp.1148-1157
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• 2006

A hyperthermophilic bacterium Thernotoga maritima produced thermostable ${\beta}-glucosidase$. The gene encoding ${\beta}-glucosidase$ from T. maritima MSB8 was cloned and expressed in Escherichia coli. The en-zyme (BgIB) hydrolyzed ${\beta}-glucosidase$ linkages between glucose and alkyl, aryl of saccharide groups such as salicin, arbutin, and $_pNPG$. The insert DNA contained ORF with 2,166 bp encodes a 721 amino acids (calculated molecular mass of 80,964 and pl of 4.93). The amino a.id sequence of BglB showed the similarity to family 3 glycosyl hydrolases. The molecular weight of the enzyme was estimated to be approximately 81kDa by MUG-nondenaturing PAGE (4-methylumbelliferyl 13-D-glucoside-nondenaturing polyacrylamide gel electophoresis) and SDS-PACE. The ${\beta}-glucosidase$ exhibited maximal activity at pH 7.0 and $80^{\circ}C$. By exchanging two possible residues (Glu-232 and Asp-242) to Ala by site-directed mutagenesis method, it was found that these were essential for enzymatic activity.

• Journal of the Korean Chemical Society
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• v.44 no.1
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• pp.87-91
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• 2000

• The Korean Journal of Food And Nutrition
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• v.11 no.6
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• pp.673-677
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• 1998

A inhibitor acting on substrate proteolytic enzyme was isolated from culture broth of Streptomyces sp. SK-862, which had been isolated from soil in Wonju City, by using the colloidal agar medium. The optimum culture temperature and initial pH for the production of the protease inhibitor was 28$^{\circ}C$ and pH 8.5, respectively. The optimum culture medium was composed of 1.5% glucose, 0.5% peptone, 0.1% K2PHO4, 0.05% CaCO3 and initial pH 8.5. The inhibitor production was maximum when the strain was incubated in shaking incubator at 70 strokes for 60 hours.

• The Korean Journal of Food And Nutrition
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• v.11 no.6
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• pp.678-682
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• 1998

A strain of Streptomyces sp. SK-862, isolated from soil in Wonju city, was able to prodce a biologically active substance that has a strong inhibitory activity against proteolsis by trypsin. The inhyibitory substance was extracted by n-butanol, and then purified by the adsorption chromatography followed by the reverse-phase high performacne liquid chromatography. The purified substance was stable over the pH range from 2 to 10, but was unstable when treated at 8$0^{\circ}C$ for 60 min. This substance was soluble in water, methanol, ethanol nd butanol, but insoluble in chlorofrom and ethylacetate. The Rf value of the purified substance on the thin layer chromatography were 0.56 in n-butanol : methanol : water(5 : 3 : 1v/v) solvent system compare dto 0.23 in ethanol : ammonium hydoxide : water(8 : 1 : 1v/v) solvent system. This substance has maximum absorption at 259 nm. The chemical reaction of the substance was negative for sugar but positive for ninhydrine and iodine reaction.

• Korean Journal of Microbiology
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• v.29 no.3
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• pp.167-171
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• 1991

An intracellular protease from cells of Pseudomonas carboxydohydrogena grown on nutrient broth was purified to better than 95% homogeneity in five steps using azocaseine as a substrate. The molecular weight of the native enzyme was determined to be 125, 000. Sodium dodecyl sulfate-gel electrophoresis revealedat least two non-identical subunits of molecular weight 70, 000 and 56, 000. The enzyme activity was completely ingibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate. The enzyme was also inhibited by $Mg^{2+}$ , $Zn^{2+}$ , $Cd^{2+}$, $Cu^{2+}$ , and $Fe^{2+}$ , but was stimulated by iodoacetamide. Maximal reaction rate of the enzyme was observed at pH8.0 and 30.deg.C. The isoelectric point of the enzyme was found to be 7.5. The enzyme was unable to hydrolyze carbon monoxide dehydrogenase.

• Journal of Pharmaceutical Investigation
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• v.19 no.2
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• pp.81-86
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• 1989

The proteolytic enzyme extracted from Neungee [Sarcodon aspratus (Berk.) S. Ito] was purified by using Tris-acryl CM-cellulose column chromatography and chromatofocusing. The specific activity of the purified enzyme increased 15.8 times as compared with that of the crude enzyme. The enzyme was homogeneous on polyacrylamide gel electrophoresis and stable at pH values ranging from 4.0 to 10.8. The enzyme activity remained unchanged when the mushroom and the purified enzyme were stored for 3 years and 6 months at 4°C, respectively. The enzyme was found to be an endogeneous protease.

• Journal of the Korean Society of Tobacco Science
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• v.16 no.1
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• pp.90-96
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• 1994

Changes in haemolymph proteins, hydrolases such as esterase(EST), acid phosphatase(ACP) and alkaline phosphatase(ALP) , and inorganic ion(Na+, K+ and Cl- ) contents were induced by the injection of Bacillus thuringiengis into haemocoel of the last instar larva of Heliothis assulta. Protein concentration of haemolymph was increased until 24 hrs after injection, and decreased thereafter. Among the 8 basic protein bands identified through acid - polyacrylamide gel electrophoresis(PAGE), 2 bands(bands a and b) became stronger by the bacterial infection. Activities of EST and ALP increased until 12 hrs after injection and then fell down, whereas ACP activity was decreased continuously with time after injection. Contents of inorganic ions were all increased by the bacterial injection, showing slow rate of increase in the chloride ion, but rapid in the sodium and potassium ions.

• The Korean Journal of Food And Nutrition
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• v.7 no.2
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• pp.128-136
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• 1994

Mackerel muscle protein hydrolysates, which were prepared from defatted mackerel meal by proteases such as complex enzyme, alcalase, bromelain, pancrease, pepsin, w-chymotrypsin, trypsin and papain, were tested for the antioxidative action against linoleic acid. Among proteases tested, the hydrolysates obtained from the treatment of complex enzyme, bromelain and alcalase showed higher antioxidative effects. Also, the hydrolysates showed the synergistic effects with o-tocopherol and the inhibitory effects for peroxidation of metal ions(Fe3+, Cua+) From the profiles of fractionation of the hydrolysates with Bio-gel P-2 column, the most active fractions, part I(complex enzyme-derived) and part e(bromelain-derived), had below MW 1,400 and the antioxidative effects were closely related to the binding capacity with metal ion(Cua+). Amno acid composition of the part I was abundant in histidine, arginine, phenylalanine and lysine, and the part e was abundant in lysine, glutamic acid and leucine.

• Korean Journal of Food Science and Technology
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• v.29 no.5
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• pp.1052-1056
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• 1997

When casein was hydrolyzed by trypsin, alcalase, neutrase, protamax, and S. aureus type V8, peptides $(100\;{\mu}g/mL)$ which were produced by trypsin and alcalase solubilized $6.42\;and\;2.37\;{\mu}g/mL)$ of added irons at pH 6, respectively, while peptides which were produced by other proteases solubilized less than $1\;{\mu}g/mL$. Peptides produced by trypsin and alcalase were fractionated to 10 fractions on a reverse phase column and each fraction was tested for its iron solubilizing ability at pH 6. Among peptides produced by trypsin, fraction 5 showed the highest iron solubilizing ability $(2.33\;{\mu}g/mL)$. In the case of alcalase, fraction 7 showed the highest iron solubilizing ability $(1.56\;{\mu}g/mL)$. To isolate iron binding peptides from peptides produced by trypsin and alcalase, immobilized iron affinity chromatography which irons were chelated to imino diacetic acids in chelating sepharose fast flow were utilized. Our results showed that immobilized iron affinity chromatography was an effective method to isolate iron binding peptides produced by either trypsin or alcalase from milk casein.

• Microbiology and Biotechnology Letters
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• v.36 no.4
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• pp.299-306
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• 2008

Human pancreatic lipase is a digestive enzyme which is synthesized in pancreas, secreted into small intestine, and there hydrolyze the fat in food. Pancreatic lipase protein composes of catalytic domain and colipase-binding domain. In this research, the gene segments corresponding to total protein, catalytic domain, and co lipase-binding domain were cloned by PCR method, inserted into an expression vector, and then used to transform Escherichia coli BL21 (DE3). The recombinant proteins produced were purified and injected intramuscularly three times into laying hens. The egg yolk antibodies (IgY) were obtained from the egg yolks and tested for their antibody titer. Among three IgY, the IgY against colipase-binding domain showed the highest antibody titer. All three IgY had inhibitory effects on the porcine pancreatic lipase. Among them, the IgY against colipase-binding domain showed the highest inhibition effects. The fat diet with corn oil and IgY was administrated to the experimental rats and their blood compositions were examined with time course. The triglyceride concentration of treated rats was decrease meaningfully when compared with those of control rats. This suggested that the IgY against colipase-binding domain antigen inhibited pancreatic lipase in vivo.