• Journal of Microbiology and Biotechnology
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• v.16 no.5
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• pp.757-763
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• 2006

Angiotensin I-converting enzyme (ACE) inhibitors have generally been very useful to remedy or prevent hypertension. This study describes the extraction and characterization of an ACE inhibitor from the fruiting body of Pholiota adiposa ASI 24012, which can be used as an antihypertensive drug. The maximal ACE inhibitory activity $(IC_{50};0.25mg)$ was obtained when the fruiting body of Pholiota adiposa ASI 24012 was extracted with distilled water at $30^{\circ}C$ for 12 h. After the purification of ACE inhibitor with ultrafiltration, Sephadex G-25 column chromatography, and reverse-phase HPLC, an active fraction with an $IC_{50}$ of 0.044 mg was obtained. The purified ACE inhibitory peptide was a novel pentapeptide, showing very little similarity to other ACE inhibitory peptide sequences. The molecular mass of the purified ACE inhibitor was estimated to be 414 daltons with a sequence of Gly-Glu-Gly-Gly-Pro, and showed a clear antihypertensive effect on spontaneously hypertensive rats (SHR) at a dosage of 1 mg/kg.

• Korean Journal of Plant Resources
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• v.22 no.3
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• pp.203-208
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• 2009

Lunasin is a unique 43-amino acid peptide which has shown a chemopreventive in mammalian cells and in a skin cancer mouse model. In search for new sources of lunasin and the role of cereals in cancer prevention, we report here the properties of lunasin purified from millet. Stability of millet lunasin was measured by in vitro digestibility assay using pepsin and pancreatin. Inhibition of HAT (histone acetyltransferase) and nuclear localization in mammalian cells were used to measure lunasin bioactivity as the cancer chemopreventive agent. Lunasin present in millet crude protein was stable to pepsin and pancreatin in in vitro digestion and inhibited the activities of HATs. When added exogenously, lunasin purified from millet internalized in the nuclei of mouse fibroblast cells. On the base of this result, we conclude that lunasin in millet is bioactive and consumption of millet may play an important role on cancer prevention in millet-consuming populations.

• Journal of Microbiology and Biotechnology
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• v.7 no.6
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• pp.402-408
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• 1997

The antibiotic-producing strain HW-003 was screened from soil and found to be effective against the multidrug-resistant Staphylococcus aureus. The spore chain of HW-003 was retinaculiaperti, and the spore surface was spiny. Strain HW-003 has a LL-diaminopimelic acid isoform in the cell wall. The aerial mass color of the strain was gray, and the reverse side was yellow-brown. The strain produced melanin, but did not produce soluble pigments. According to the Taxon program, HW-003 showed best match with Streptomyces cyaneus. Antibiotic production reached a maximum after 72-h cultivation. The antibiotic was purified with silica gel column chromatography, octadecylsilyl column chromatography, and HPLC. The purified antibiotic, AMRSA1, showed strong inhibitory activity against multidrug-resistant Staphylococcus aureus and gram-positive bacteria. The molecular weight of AMRSA1 was about 1, 100. AMRSA1 was a peptide antibiotic containing alanine and serine.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.28 no.6
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• pp.728-735
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• 1995

The peptidyl prolyl cis-trans isomerase(PPlase, EC 5.2.2.8) from Bacillus stearothermophilus SIC1 was extracted from the cells treated with by lysozyme. PPlase was purified from the cell extracts by heat treatment, ammonium sulfate precipitation, ion exchange chromatography and finally gel filtration (FPLC). The purity of purified the enzyme after Superose 12 column chromatography was examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE). The molecular weight of the purified PPlase was estimated as 18,000 by SDS-PAGE. The 39 amino acid residues from the N-terminus were determined by the protein sequencer. The enzyme showed the optimum pH at 8.0 and was stable at the range of pH 7.0 to 8.0. The enzyme was considerably stable after heat treatment at $60^{\circ}C$ for 30 minutes, and the enzyme was quite stable up to $65^{\circ}C$. The presence of the PPlase in the refolding solution accelerated the isomerization rate of the assay peptide.

• Preventive Nutrition and Food Science
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• v.5 no.2
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• pp.75-80
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• 2000

To separate ACE inhibitors from edible plants, spices, and herbs, 285 extracts of 95 sources were screened for ACE inhibitory activity. The extract of Sinapis alba L. had the most potent ACE inhibitory activity. Mustard seeds were crushed homogeneously and extracted with hexane and water successively. Lyophilized water extract was fractionated with $H_2O$:butanol(1:1). The ACE inhibitor was purified from butanol fraction by methanol precipi-tation, gel filtration, HPLC, and FPLC with Superdex peptide HQ 10/30 column. The active fraction has been purified to homogeneity, which was proven by gel filtration using FPLC system. The yield was 0.02%. The com-pound has a molecular weight of about 640. The compound competitively inhibited ACE activity and the $IC_{50}$ value was 79$\mu\textrm{g}$/ml. The purified compound showed uterus contraction activity in isolated rat uterus.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.38 no.1
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• pp.6-11
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• 2005

Vasopressin (VP)-related peptide was purified from the brain extract of conger eel (Conger myriaster) by reverse-phase, ion-exchange high performance liquid chromatography (HPLC). This peptide with a molecular weight of 1,051.2 Da was determined as $H-Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Arg-Gly-NH_2$, whose Cys residues made an intramolecular disulfide bridge by the automated amino acid sequence analysis, MALDI- TOF mass spectrometry. It's sequence was confirmed by identity of the elution position with the synthetic peptide in HPLC system. As a result of homology investigation, the primary structure of this peptide was the same as that of VP-superfamily member, $[Arg^8]-vasotocin$. The synthetic peptide showed a contractile activity at a minimal effective concentration of $10^{-10}\;M$ on the intestinal smooth muscle of goldfish.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.8
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• pp.1124-1129
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• 2005

Functional activities of low molecular weight substances purified from pepsin hydrolysates of four different seaweeds; Costaria costata, Enteromorpha prolifera, Grateloupia filicina and Porphyra tenera, were inves-tigated. Each pepsin hydrolysate of Costaria costata, Enteromorpha prolifera, and Grateloupia filicina resulted in three peptide peaks on Bio-Rad P2 gel chromatography pattern, while that of Porphyra tenera showed 2 peaks. Peak 1 of Porphyra tenera showed the highest antioxidative activity followed by peak 2 of Porphyra tenera and peak 2 of Costaria costata in order Peak 1 of Porphyra tenera showed the highest ACE inhibitory activity followed by peak 3 and peak 2 of Enteromorpha prolifera in order. Peak 1 and peak 2 of Porphyra tenera, and peak 2 of Enteromorpha prolifera showed the highest antityrosinase activity followed by peak 3 of Enteromorpha prolifera. Peak 1 of Enteromorpha prolifera showed the highest antitumor activity followed by peak 2 of Costaria costata, peak 3 of Enteromorpha prolifera, and peak 3 of Grateloupia filicina in order. Porphyra tenera showed the highest functional activities, which is thought to be due to its high protein content. Structure and amino acid sequence of low molecular weight peptide of Porphyra tenera should be analyzed in the further study.

• Korean Journal of Food Science and Technology
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• v.37 no.5
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• pp.827-832
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• 2005

Bacillus subtilis JM3 protease from naturally fermented anchovy sauce was partially purified in 40-60% ammonium sulfate fraction. To accelerate fermentation of anchovy sauce, 2 and 4% crude B. subtilis JM3 proteases were added to 6 month-ripened anchovy sauce, and their hydrolysis degrees and amino-nitrogen contents were investigated at different storage times. Low molecular weight (LMW) peptide was purified by ultrafiltration ana gel permeation chromatography from anchovy sauce manufactured with B, subtilis JM3 protease. Anchovy sauces with 2 and 4% proteases increased hydrolysis rate by 27 and 32%, respectively. Amino-nitrogen contents of anchovy sauces fermented with 2 and 4% proteases were twofold higher than that of control. Control showed five peptide peaks on Bio-Rad P2 gel permeation chromatography spectrum, whereas anchovy sauces with 2 and 4% B. subtilis JM3 proteases showed six and seven peaks, respectively. ACE inhibitory activity was highest in peak 6 (43.75%) of anchovy sauce with 2% protease, followed by peak 5 (34.82%) of control. DPPH radical-scavenging effect was higher than 50% in all samples. Cytotoxicity was highest in peak 3 (44.12%) of control, fellowed by peak 5 (42.04%) of anchovy sauce with 4% protease.

• International Journal of Industrial Entomology and Biomaterials
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• v.17 no.1
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• pp.137-141
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• 2008

We previously isolated and cloned a cDNA of abaecin from the Bombus ignitus. In an effort to produce a large amount of soluble abaecin at low cost, we successfully expressed the peptide in Escherichia coli that are highly sensitive to its mature form. For this, we fused the peptide encoding 39 amino acids of mature B. ignitus abaecin to the thioredoxin gene together with a C-terminal 6xHis tag. An enterokinase cleavage site was introduced between the 6xHis tag and mature abaecin to allow final release of the recombinant peptide. A high yield of 9.6 mg soluble fusion protein from 200 ml of bacterial culture was purified by $Ni^{2+}$-charged His-Bind resin affinity column, and 1.4 mg of pure active recombinant abaecin was readily obtained by enterokinase cleavage, followed by affinity chromatograph. The molecular mass of recombinant abaecin peptide was determined by Tricin-SDS-PAGE analysis. The recombinant abaecin exhibited antibacterial activity against Gram-negative bacteria.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1620-1627
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• 2009

The design and expression of an antihypertensive peptide multimer (AHPM), a common precursor of 11 kinds of antihypertensive peptides (AHPs) tandemly linked up according to the restriction sites of gastrointestinal proteases, were explored. The DNA fragment encoding the AHPM was chemically synthesized and cloned into expression vector pGEX-3X. After an optimum induction with IPTG, the recombinant AHPM fused with glutathione S-transferase (GST-AHPM) was expressed mostly as inclusion body in Escherichia coli BL21 and reached the maximal production, 35% of total intracellular protein. The inclusion body was washed, dissolved, and purified by cation-exchange chromatography under denaturing conditions, followed by refolding together with size-exclusion chromatography and gradual dialysis. The resulting yield of the soluble GSTAHPM (34 kDa) with a purity of 95% reached 399 mg/l culture. The release of high active fragments from the AHPM was confirmed by the simulated gastrointestinal digestion. The results suggest that the design strategy and production method of the AHPM will be useful to obtain a large quantity of recombinant AHPs at a low cost.