• Journal of Ginseng Research
• /
• v.19 no.3
• /
• pp.237-243
• /
• 1995

Agmatine iminohydrolase (EC 3.5.3.12) catalyzes the hydrolysis of agmatine into putrescine. The enzyme seems to be one of the critical enzymes in putrescine biosynthesis. The enzyme was purified to homogeneity from Panax ginseng C.A. Meyer by combined method of ammonium sulfate 1 fractionation, DEAR anion exchange column, hydroxyapatite column and agmatine carboxyhexyl Sepharose 4B affinity column. The molecular weight estimated by native pore gadient polyacrylamide gel electrophoresis was 71, 000 Dalton, while that estimated by SDS-PAGE was 70, 000 Dalton, indicating a monomeric enzyme. The optimal pH and temperature were 9.0 and 37$^{\circ}C$, respectively. The Km and 1 Vmax for agmatine were 8.3 mM and 14.4 nmole/hr, respectively. Heat stability of this enzyme was high. The enzyme was observed to be inhibited by polyamines such as putrescine, cadaverine, spermidine and spermine. Especially, putrescine was a potent inhibitor of the purified enzyme. These results suggest that polyamines could be important in growth regulation of Panax ginseng C.A. Meyer.

• The Journal of Natural Sciences
• /
• v.14 no.2
• /
• pp.55-65
• /
• 2004

A thiol-specific antioxidant(TSA) protein was purified from Earthworm, Lumbricus terrestris by DEAE-Cellulose, Phenl sepharose, Sephacryl S-200 gel filtration and HPLC S-300 Column Chromatography. This protein showed a thiol-specific antioxidant activity against inactivation of glutamine synthetase by a metal-catalysed oxidation system capable of generation reaction oxygen species. The molecular mass of the protein was determinated to be 51-kDa by SDS-polyacrylamide gel electrophores. Taken together, the purified TSA protein could be a new member of TSA family.

• Journal of Aquaculture
• /
• v.10 no.4
• /
• pp.425-433
• /
• 1997

The isolated soluble eye proteins from six species (Sepia esculenta, Sepiella japonica, Loligo chinensis, Todarodes pacificus, Loligo beka and Ocotopus minor) of class Cephaloopoda were compared by crossed immunoelectrophoresis methods using antibodies directed against total soluble eye proteins antigens. It showed a distinct antigen-antibody reaction between the species of order Sepioidea and various antiserum of class Cephalopoda. Our results suggested that the common precipitation lines of Sepia and Loligo species which were different from that of Octopus minor. By which obtained from elution of gel filtration chromatography and 12.5% SDS-polyacrylamide gel electrophoresis, the molecular weight of Todarodes pacificus eye protein (crystallin) was determined in abut 20-35 KDa.

• The Korean Journal of Zoology
• /
• v.22 no.4
• /
• pp.141-152
• /
• 1979

Feline leukemia virus DNA polymerase was purified by ion-exchange and nucleic acid affinity chromatographies. The enzyme consists of a single polypeptide chain of approximately 72, 000 molecular weight as determined by both of a glycerol density gradient centrifugation and SDS-polyacrylamide gel electrophoresis. The preferred divalent cation for DNA synthesis is $Mn^2+$ on a variety of template-primers, and its optimum concentration appears to be significantly lower than reported results of other mammalian type-C viral enzymes. The divalent cation requirement for maximum activity of RNase H is similar to those of DNA polymerase. Both DNA polymerase and RNase H activities appear to reside on the same molecule as demonstrated by the copurification of both activities through various purification steps. An additional RNase H without detectible polymerase activity was generated by a limited chymotrypsin digestion. This RNase H activity was inhibited equally effectively as RNase H in the intact reverse transcriptase by antisera prepared against reverse transcriptase of feline leukemia virus. Neutralization and binding test showed that antibody binding to reverse transcriptase molecule did not completely inhibit the polymerase activity.

• Archives of Pharmacal Research
• /
• v.19 no.3
• /
• pp.191-196
• /
• 1996

Bizelesin is a promising novel anticancer agent which is known to alkylate N3 of adenine to induce DNA interstrand cross-links (ISC) with in $5^I-TAATTA\; and\; 5^I-TAAAAAA$. We have investigated the base specificity for DNA ISC induced by bizelesin using oligomers containing the cross-linkable sequence $5^I-TAATTA\; and\; 5^I-TAAAAAA$. in which "N" was either A, C, G, or T. An analysis of denaturing polyacrylamide gel showed that bizelesin is able to induce DNA ISC in the duplex oligomer containing sequences $5^I-TAATTA\; and\; 5^I-TAAAAAA$. The formation of interstrand crosslinking did not occur in the sequences $5^I-TAATTA\; and\; 5^I-TAAAAAA$. DNA strand cleavage assay to determine the cross-linking site within $5^I-TAATTA$sequence showed that bizelesin alkylates guanine. These results demonstrate that bizelesin is able to induce DNA ISC at guanine but not at cytosine or thymine. In addition, guanine adducts have been found to be susceptible to DNA strand cleavage by exposure to hot piperidine. The extent of DNA strand cleavage, however, was not 100% efficient in either neutral pH buffer or hot piperidine.

• YAKHAK HOEJI
• /
• v.49 no.2
• /
• pp.122-127
• /
• 2005

Dental caries is one of the most common oral diseases in the world. Glucosyltransferase (Gtase) of Streptococcus mutans (S. mutans) plays an important role in the develo pment of dental caries. For the purpose to develop anti- caries, we examined the effect of metabolites isolated from Streptomyces sp. M-20 on Gtase and the growth of S. mutans. Streptomyces sp. M-20 isolated from Mongolian soil showed 95~96% sequence homology with that of Streptomyces lin- colnensis. The metabolites of Streptomyces sp. M-20 were partially purified by extraction with ethyl acetate, silica gel column chromatography and preparative TLC. Partially purified metabolite, red colored component (MR-20) in ethyl acetate fraction showed potent antibacterial activitiy against S. mutans and inhibitory activity against Gtase purified from S. mutans, while another isolated yellow component (MY-20) showed no activity against S. mutans. The inhibitory activity of MR-20 against Gtase was confirmed by activity staining on SDS-polyacrylamide gel electrophoresis. The concentration of MR-20 for 50% inhibition $(IC_{50})$ against Gtase activity was $60{\mu}g/ml$. These results suggest that MR-20 can be developed for antibacterial agent and anticaries.

• Journal of Life Science
• /
• v.16 no.7 s.80
• /
• pp.1225-1228
• /
• 2006

Cyclic AMP receptor protein (CRP) complexed with cAMP binds to DNA and induces sharp DNA bending around ${\sim}90$ degree. Previous publication (5), however, reported that mutant CRP:cGMP complex showed high migration rate relative to mutant CRP:cAMP complex on native polyacrylamide gel. To confirm DNA structural change in the presence of CRP and cyclic nucleotide, molar cyclization factor $(j_M)$ [13] was measured with 6 constructed DNA fragments. Nonlinear regression analysis of $j_M$ data indicated that mutant CRP did not induce DNA bending in the presence of cGMP but bent DNA in the presence of cAMP without any helical twist change in DNA.

• Journal of Pharmacopuncture
• /
• v.14 no.3
• /
• pp.71-79
• /
• 2011

Objectives : This study was undertaken to identify fibrinolytic proteases from Gloydius blomhoffii siniticus venom and to characterize a major fibrinolytic protease purified from the venom. Methods : The venom was subjected to chromatography using columns of Q-Sepharose and Sephadex G-75. The molecular weights of fibrinolytic proteases showing fibrinolytic zone in fibrin plate assay were determined in SDS-PAGE (Sodium dodecyl sulfate-polyacrylamide gel electrophoresis) The effects of inhibitors and metal ions on fibrinolytic protease and the proteolysis patterns of fibrinogen, gelatin, and bovine serum albumin were investigated. Results : 1) The fibrinolytic fractions of the three peaks isolated from Gloydius blomhoffii siniticus venom contained two polypeptides of 46 and 59 kDa and three polypeptides of 32, 18, and 15 kDa and a major polypeptide of 54 kDa, respectively. 2) The fibrinolytic activity of the purified protease of 54 kDA was inhibited by metal chelators, such as EDTA, EGTA, and 1,10-phenanthroline, and disulfhydryl-reducing compounds, such as dithiothreitol and cysteine. 3) Calcium chloride promoted the fibrinolytic activity of the protease, but mercuric chloride and cobalt(II) chloride inhibited it. 4) The fibrinolytic protease cleaved preferentially A${\alpha}$-chain and slowly B${\beta}$-chain of fibrinogen. It also hydrolyzed gelatin but not bovine serum albumin. Conclusions : The Gloydius blomhoffii siniticus venom contained more than three fibrinolytic proteases. The major fibrinolytic protease was a metalloprotease which hydrolyzed both fibrinogen and gelatin, but not bovine serum albumin.

• Korean Journal of Microbiology
• /
• v.27 no.4
• /
• pp.323-333
• /
• 1989

Plasmid DNA molecules containing strong promoter upstream from IS50L or IS50R, the two insertion sequences that flank Tn5, were constructed to amplify the synthesis of Tn5-encoded polypeptides. When proteins made by cells that contain these plasmids were analyzed on polyacrylamide gels, enhanced synthesis of IS50R polypeptides could be detected. Synthesis of this polypeptide apparently is initiated within the large open reading frame of this element. In addition, the stability of IS50L-and IS50R-encoded polypeptides was analyzed. It was found that IS50L polypeptides are relatively unstable in vivo. This instability could account for the observed inability of this element to promote transposition.

• Journal of Microbiology
• /
• v.37 no.1
• /
• pp.21-26
• /
• 1999

Two novel calcium-binding proteins, named CAB-I and CAB-II, have been isolated from Streptomyces coelicolor. Purification of the calcium-binding proteins involved heat treatment, fractionation with ammonium sulfate, acid treatment, anion exchange and hydrophobic interaction column chromatography, FPLC gel filtration, and preparative isoelectric focusing. A chelex competitive assay and 45Ca autoradiography verified the calcium-binding ability of the proteins. The major band CAB-II has an apparent molecular weight of 26,000 determined by SDS-polyacrylamide gel electrophoresis and 340,000 determined by gel filtration. The isoelectric point of this molecule showed the acidic nature of the molecule. N-terminal amino acid sequence analysis shows homology to rat Ca2+/calmodulin-dependent protein kinase-II (CAB-II) and yeast phosphoprotein phosphatase (CAB-I).