• Journal of the Korean Chemical Society
• /
• v.31 no.5
• /
• pp.457-463
• /
• 1987

The total synthesis of insulin A chain (1-21) with properly protected sulfhdryl groups of three cysteins for the correct intra and inter disulfide bond formation has been accomplished on 2-bromopropionylated 2% DVB-styreneresin support employing manually operated rotary vessel. The sulfhydryl groups of cysteins were protected with acetamidomethyl, benzyl, and benzhydryl respectively. Glutamine and asparagine were attached to the peptide chain by active ester coupling, all other amino acids were coupled with DCC/HOBT. The synthesized peptide was purified by DEAE Sephadex A-25 and gel filtration Sephadex LH-20. The final product was found to be homogeneous by HPLC, electrophoresis, and amino acid analysis. The overall yield of the pure isolated peptide was 6%.

• Radiation Oncology Journal
• /
• v.8 no.2
• /
• pp.145-150
• /
• 1990

Mesna has been used with ifosfamide to prevent urotoxicity in the treatment of testicular cancers. This drug also protected the toxicities of adriamycin without compromising cytostatic activity. With an idea of radioprotective role of sulfhydryl group of radioprotectors and of mesna decreasing the toxic effect of adriamycin which produces free radicals, mesna and radiation were administered to mice to study the protective effect of this drug and to identify the difference in regenerative capacity of the germ cells in the testis between radiation-treated and both mesna-and radiation-treated groups. The shape and numbers of spermatogenic cells in the seminiferous tubules were examined every week after irradiation. In both groups, initial reduction and later recovery in germ cell numbers and shape was observed. The lowest germ cell number was found around three weeks after irradiation. Mean germ cell number of the mesna-treated group was significantly higher than radiation-treated group at all observed periods (p<0.05). More competent regeneration was present in mesna-treated group. These results suggest that mesna protect the testis from radiation injury. Further study will be necessary to identify whether mesna protects other tissues from radiation and it does not hamper tumor control.

• Asian-Australasian Journal of Animal Sciences
• /
• v.18 no.8
• /
• pp.1130-1134
• /
• 2005

An experiment was performed using 20 calves of about one-month old to investigate the effect of chelation therapy with calcium disodium ethylenediaminetetraacetate ($CaNa_2$EDTA) alone or along with antioxidant $\alpha$-tocopherol in lead loaded calves on blood trace minerals, erythrocytic sulfahydryl groups and some haematobiochemical parameters. Fifteen calves were given lead orally at a daily dose of 7.5 mg of 99% pure lead acetate/kg body weight for 28 days. Then the lead was withdrawn on day 28 and the calves were randomly divided into three groups. Each group of five animals was either treated with $CaNa_2$EDTA alone at the dose rate of 110 mg/kg body weight in two divided doses for 4 days or along with $\alpha$-tocopherol at the dose rate of 100 mg/kg body weight orally daily for 7 days, keeping the remaining five calves as lead-exposed untreated controls. Blood samples were collected at the end of the lead exposure (day 0) and thereafter on day 2, 4, 7 and 10 from the start of the chelation treatment. The treatment with EDTA alone led to slow but non-significant improvement in blood copper level, but incorporation of antioxidant $\alpha$-tocopherol in chelation therapy resulted in its significant decline, as recorded on day 7-post treatment. Withdrawal of lead or treatment with $CaNa_2$EDTA alone or along with $\alpha$-tocopherol enhanced the erythrocytic thiol contents and the levels of T-SH and P-SH became statistically (p<0.05) comparable to those of lead-exposed controls by day 7 and 4, respectively. There was no significant (p>0.05) change in serum urea, creatinine, total protein and albumin levels between the treatment groups. It is concluded from the present investigation that treatment with $CaNa_2$EDTA at the present dose rate is safe to be used for chelation in lead loaded calves.

• Molecules and Cells
• /
• v.20 no.1
• /
• pp.127-135
• /
• 2005

The uptake of putrescine, spermidine and spermine by Fortner's hamster amelanocytic melanoma AMEL-3 cells was observed in this study to be time-dependent, temperature-sensitive, pH-dependent and saturable. Metabolic poisons nullified polyamine uptake, an indication that this is an energy-requiring mechanism. The presence of $Na^+$ ions was found to be requisite to full activity. Valinomycin, gramicidin, monensin and the calcium ionophore calcimycin were also observed to inhibit the process substantially. The transporter active site would seem to contain sulfhydryl groups. Other diamines and polyamine analogues, as well as cationic diamidines, suppressed putrescine uptake. The presence of the ornithine decarboxylase inhibitor DFMO in the culture medium induced putrescine inflows. Putrescine, in turn, induced the negative expression of the carrier, thus suggesting that this influx mechanism is governed by up/down regulation. The cationic diamidine CGP 40215A and its analogue CGP039937A competitively inhibited putrescine transport, with Ki values of 1.9 and $15{\mu}M$, respectively. The role of polyamine uptake in these cultures is discussed.

• Journal of Microbiology and Biotechnology
• /
• v.8 no.6
• /
• pp.700-704
• /
• 1998

Each of the recombinant structural genes of Helicobacter pylori urease, ureA and ureB, was cloned and overexpressed as inclusion bodies. Solubilization and renaturation of the inclusion bodies were carried out, to accelerate the pairing of sulfhydryl groups and the incorporation of nickel ions, which would lead to the native structure with high enzyme activity. Rates of urea hydrolysis were monitored as an indication of in vitro activation of renatured ureases. The activation of the apoprotein using 1 mM nickel ion, 100 mM sodium bicarbonate and a 10:1 ratio of reducing power resulted in a weak urease activity (about 11% of the native urease activity encoded by pTZ 19R/ure-l). When a sparse matrix screen method originally discovered for the crystallization of proteins was used, the activity increased higher than that obtained using glutathione. The effect of polyethylene glycol (PEG) on the activity was noticeable, giving two-fold increase in the specific activity (about 11 U/mg of protein corresponding to 22% of the native urease activity encoded by pTZ19R/ure-1).

• Journal of Pharmaceutical Investigation
• /
• v.41 no.2
• /
• pp.91-94
• /
• 2011

The purpose of this study was to investigate the effect of peptide charge on the interaction between peptide and poly(D,L-lactide-co-glycolide) (PLGA) for evaluating mechanism of acylated peptide formation in PLGA matrix. As a model peptide, octreotide, a synthetic somatostatin analogue and active ingredient of commercial PLGA product, was used. The disulfide group of octreotide was reduced with dithiothreitol and the sulfhydryl groups were modified with N-${\beta}$-maleimidopropionic acid (BMPA) to neutralize octreotide with positive charge in physiological conditions. The BMPA-conjugated octreotide was identified by measuring the molecular mass with liquid chromatography-mass spectrometry. In the interaction study with PLGA, native octreotide showed initial adsorption to PLGA and substantial production of acylated peptides (56% of overall peptide), whereas BMPA-conjugated octreotide showed minimal adsorption to PLGA and no acylation products for 42 days. Consequently, the neutralization of octreotide completely inhibited the peptide acylation by preventing interaction of peptide with PLGA. In conclusion, this study demonstrates that the initial polymer interaction of peptide is important step for peptide acylation in PLGA matrix and suggests the modulation of peptide charge as strategy for inhibiting the formation of acylated peptide impurities.

• BMB Reports
• /
• v.36 no.2
• /
• pp.149-153
• /
• 2003

The dibucaine number (DN) was determined for serum cholinesterase (EC 3.1.1.8, SChE) in plasma samples. The ones with a DN of 79-82 were used, because they had the "usual" SChE variant. The enzyme was assayed colorimetrically by the reaction of 5,5'-dithiobis-[2-nitrobenzoic acid] (DTNB) with the free sulfhydryl groups of thiocholine that were produced by the enzyme reaction with butrylthiocholine (BuTch) or acetylthiocholine (AcTch) substrates, and measured at 412 nm. Dibucaine, a quaternary ammonium compound, inhibited SChE to a minimum within 2 min in a reversible manner. The inhibition was very potent. It had an $IC_{50}$ of $5.3\;{\mu}M$ with BuTch or $3.8\;{\mu}M$ with AcTch. The inhibition was competitive with respect to BuTch with a $K_i$ of $1.3\;{\mu}M$ and a linear-mixed type (competitive/noncompetitive) with respect to AcTch with inhibition constants, $K_i$ and $K_I$ of 0.66 and $2.5\;{\mu}M$, respectively. Dibucaine possesses a butoxy side chain that is similar to the butryl group of BuTch and longer by an ethylene group from AcTch. This may account for the difference in inhibition behavior. It may also suggest the existence of an additional binding site, other than the anionic binding site, and of a hydrophobic nature.

• BMB Reports
• /
• v.37 no.5
• /
• pp.597-602
• /
• 2004

N-terminal His-tagged recombinant $\beta$-1,4-galactosyltransferase from Neisseria meningitidis was expressed and purified to homogeneity by column chromatography using Ni-NTA resin. Mutations were introduced to investigate the roles of, Ser68, His69, Glu88, Asp90, and Tyr156, which are components of a highly conserved region in recombinant $\beta$-1,4 galactosyltransferase. Also, the functions of three other cysteine residues, Cys65, Cys139, and Cys205, were investigated using site-directed mutagenesis to determine the location of the disulfide bond and the role of the sulfhydryl groups. Purified mutant galactosyltransferases, His69Phe, Glu88Gln and Asp90Asn completely shut down wild-type galactosyltransferase activity (1-3%). Also, Ser68Ala showed much lower activity than wild-type galactosyltransferase (19%). However, only the substitution of Tyr156Phe resulted in a slight reduction in galactosyltransferase activity (90%). The enzyme was found to remain active when the cysteine residues at positions 139 and 205 were replaced separately with serine. However, enzyme reactivity was found to be markedly reduced when Cys65 was replaced with serine (27%). These results indicate that conserved amino acids such as Cys65, Ser68, His69, Glu88, and Asp90 may be involved in the binding of substrates or in the catalysis of the galactosyltransferase reaction.

• Food Science and Biotechnology
• /
• v.16 no.6
• /
• pp.1051-1054
• /
• 2007

An edible wild perennial plant with extremely potent antimicrobial activity was found and identified as apiifolia Virgin's Bower (Clematis apiifolia DC) which is easily found around wet wildernesses. Fresh fruit extract of C. apiifolia exhibited minimum inhibitory concentrations (MIC) in the vicinity of 0.1% against various yeasts and of less than or equal to 0.4% for non-lactic acid bacteria. MICs against lactic acid bacteria were about 2.0%. The antimicrobial activity of C. apiifolia fruit was even more potent than that of garlic which has been known for its potent antimicrobial activity. The principal antimicrobial compound of fruit extract of C. apiifolia was isolated and identified by high performance liquid chromatography and gas chromatography as protoanemonin (a gamma lactone of 4-hydroxy-2,4-pentadienoic acid). The antimicrobial activity of C. apiifolia was stable at high temperatures, and the activity was maintained after heating at $121^{\circ}C$ for 10 min. The antimicrobial compound of C. apiifolia was supposed to inhibit microorganisms by reacting with sulfhydryl groups of cellular proteins.

• Food Science of Animal Resources
• /
• v.21 no.1
• /
• pp.56-63
• /
• 2001

In this study, three domestic and one foreign formulas for the infants up to 5 month old were examined to detect chemical changes such as pH, reactive sulfhydryl groups(RSH) content, 5-hydroxymethylfurfral(HMF) content, available lysine content, electrophoresis, and surface color caused by heat treatment for long term storage. In the SDS-polyacrylamide gel electrophoresis, A and B products showed similar pattern, while C product had a clearly distinguishable $\beta$-lacto-globulin band, but in casein, only D product showed a few strong casein band. RSH content, which indicate the extent of whey protein denaturation, ranged from 4.40 to 5.93 mmole/g protein. HMF content. which indicate the extent of Maillard reaction, ranged from 192 to 432 $\mu$mole/100g in formulas. B product showed the highest RSH and HMF content. Available lysine content ranged from 31 to 46 mg/g protein. Among them D product contain the highest available lysine content and others showed no significant difference. In conclusion, the domestic infant formulas showed higher RSH and HMF content than the foreign product and the available lysine content of the domestic products were lower than of the foreign product.