• Asian-Australasian Journal of Animal Sciences
• /
• 제25권6호
• /
• pp.852-860
• /
• 2012

An Antarctic bacterial isolate displaying extracellular ${\alpha}$-galactosidic activity was named Paenibacillus sp. LX-20 based on 16S rRNA gene sequence analysis. Optimal activity for the LX-20 ${\alpha}$-galactosidase occurred at pH 6.0-6.5 and $45^{\circ}C$. The enzyme immobilized on the smart polymer Eudragit L-100 retained 70% of its original activity after incubation for 30 min at $50^{\circ}C$, while the free enzyme retained 58% of activity. The enzyme had relatively high specificity for ${\alpha}$-D-galactosides such as p-nitrophenyl-${\alpha}$-galactopyranoside, melibiose, raffinose and stachyose, and was resistant to some proteases such as trypsin, pancreatin and pronase. Enzyme activity was almost completely inhibited by $Ag^+$, $Hg^{2+}$, $Cu^{2+}$, and sodium dodecyl sulfate, but activity was not affected by ${\beta}$-mercaptoethanol or EDTA. LX-20 ${\alpha}$-galactosidase may be potentially useful as an additive for soybean processing in the feed industry.

• 한국미생물·생명공학회지
• /
• 제18권4호
• /
• pp.360-367
• /
• 1990

토양으로부터 rutin을 유일한 탄소원으로 생육하는 세균을 분리하여 그 중 rutin 가수분해활성이 높은 MT-57균을 Arthrobacter sp.로 동정하였다. Arthrobacter sp.MT-57에서 수 종류의 rutin 가수분해효소가 확인되었으며 이들 중 rutinosidase를 전기영동상 단일한 효소 단백질까지 정제하였다. Rutinosidase으 분자량은 SDS page와 gel filtration으로 부터 42,000의 단량체로 추정 되었다. 효소반응의 최적 pH와 온도는 pH 7.5와 $45^{\circ}C$였으며, 20분간 열처리하였을 때 $50^{\circ}C$까지 안정하였다.

• 한국표면공학회지
• /
• 제23권1호
• /
• pp.34-43
• /
• 1990

Al-Si piston alloys such as AlS10CuMg have been anodized to examine apossibility of forming a hard film aat relatively higher temperatures compard with those in conventional sulfuric acid processes. Three types of electrolytes have been employed in this study ; electrolyte A(15% H2SO4, $0^{\circ}C$), electrolyte B(12% H2SO4, 1% oxalic, $10^{\circ}C$), electrolyte C(tartaric acid 125g/L+oxalic 75g/L+aluminum sulfate 225g/L, $25^{\circ}C$). Hard anodisine process in electrolyte B at a current density of 1.54A/dm2 produced a harder film of VHN 396 at a relatibely low film forming voltage compared with those obtained in other electrolyte at equivalent current density. A liner relationship between hardness and abrasion resistance exists for Al-Si piston alloys. The hardness of anodized film decreasees with increasing silicon content in Al-Si alloys and also with bath temperature. The film hardeness of Na-modified alloy os higher than that of P-modified alloy due to its finer microstructre. The film on the silicon phase in Al-Si alloys is observed to be formed by lateral growth of oxide film nucleated at surroundings.

• Journal of Pharmaceutical Investigation
• /
• 제19권1호
• /
• pp.9-14
• /
• 1989

In order to study the protease from Agaricus bisporus (Lange), the crude protease preparation was separated by fractionation of mushroom extracts with ammonium sulfate. It was found that extracts from Agaricus bisporus (Lange) Sing. contained protease. The optimum pH of the enzyme was 6.0, and the pH range at which the enzyme was stable was 4.0 to 7.0. The optimum temperature at which the enzyme showed the highest proteolytic activity was $50^{\circ}C$, while the enzyme was instantly inactivated at about $60^{\circ}C$. The enzyme activity was inhibited by $Ag^+$, $Hg^{2+}$, $Cu^{2+}$, $Ba^{2+}$, $Fe^{3+}$, $Co^{2+}$, $Ca^{2+}$, $Pb^{2+}$, $Mg^{2+}$ and $Mn^{2+}$. The $K_m$ value was 0.32 mM with Hammarsten casein.

• 한국미생물·생명공학회지
• /
• 제24권2호
• /
• pp.197-205
• /
• 1996

The $\beta$-xylosidase was purified 99- fold from the culture supernatant of Pseudo onas sp. CB-33 by ammonium sulfate precipitation, PEI precipita- tion, DEAE-Sephadex column chromatography, Sephadex G-75 gel filtration chromatography and preparative disc gel electrophoresis. Molecular weight of the enzyme was estimated to be 44,000 by SDS polyacrylamide gel electrophoresis. The enzyme has a pH optimum for activity at 7.0 and is stable over pH 6.5-9.0. The optimal temperature of the enzyme was 45$\circ$C, and its enzymatic activity was completely inactivated at 55$\circ$C for 30 min. Km value of the enzyme for p-nitrophenyl-$\beta$-D-xylopyranoside was calculated to be 4.6 mM. The effect of various reagents on the $\beta$-xylosidase activity was investigated. The enzyme activity was completely inhibited by Hg$^{2+}$, Cu$^{2+}$ and Zn$^{2+}$. The $\beta$-xylosidase was inactivated by tryptophan-specific reagent, N-bromosuccinimide and tyrosine-specific reagent, iodine. The enzyme could degrade xylo-oligosaccharides to xylose and the enzyme was competitively inhibited by xylose. The $\beta$-xylosidase and endoxylanase from Psedomonas sp. CB-33 hydrolized xylan synergically. The purified enzyme also showed $\alpha$-L-arabinofuranosidase activity.

• Mycobiology
• /
• 제42권4호
• /
• pp.322-330
• /
• 2014

The aim of this study was to identify and characterize new Flammulina velutipes laccases from its whole-genome sequence. Of the 15 putative laccase genes detected in the F. velutipes genome, four new laccase genes (fvLac-1, fvLac-2, fvLac3, and fvLac-4) were found to contain four complete copper-binding regions (ten histidine residues and one cysteine residue) and four cysteine residues involved in forming disulfide bridges, fvLac-1, fvLac-2, fvLac3, and fvLac-4, encoding proteins consisting of 516, 518, 515, and 533 amino acid residues, respectively. Potential N-glycosylation sites (Asn-Xaa-Ser/Thr) were identified in the cDNA sequence of fvLac-1 (Asn-454), fvLac-2 (Asn-437 and Asn-455), fvLac-3 (Asn-111 and Asn-237), and fvLac4 (Asn-402 and Asn-457). In addition, the first 19~20 amino acid residues of these proteins were predicted to comprise signal peptides. Laccase activity assays and reverse transcription polymerase chain reaction analyses clearly reveal that $CuSO_4$ affects the induction and the transcription level of these laccase genes.

• 미생물학회지
• /
• 제31권1호
• /
• pp.54-62
• /
• 1993

Intracellular adenosine deaminase (ADA) from Aspergillus oryzae was purified using ammonium sulfate fractionation, a DEAE-Sephadex A-50 anion exchange chromatography, an ultrafiltration using a PM 10 membrane and two times of Sephadex G-100 gel filtration chromatography. The enzyme was purified 151 fold with a 9% recovery. Purified enzyme gave a single protein band with a molecular weight of 105,000 delton. The enzyme was reasonably stable. The enzyme activity was kept even after 1 hr incubation at 55.deg.C, but decreased significantly at 60.deg.C. The pH optimum was found to be from 6.5 to 7.5. Among tested compounds, the substrate activity was found with adenosine, adenine arainofuranoside, formymcin A, 2'-deoxyadenosine, 3'-deoxyadenosine, 2', 3'-isopropylidene adenosine, 2,6-diaminopurine deoxyriboside, .betha.-nicotinamide adenine dinucleotide (reduced form), 6-chloropurine riboside, 2'-adenine monophosphate (AMP), 3'-AMP and 5'-AMP. The values of Km of adenosine and 2'-deoxyadenosine were calculated to be 500 and .$710\mu$m, respectively. ADA was sensitivite to $Zn^{2+}$, $^Cu{2+}$ and $Fe^{3+}$, p-chloromercuribenzoate and mersalyl acid inactivated the enzyme. The activity of enzyme was not changed when ADA was incubated with dithiothreititol, 2-mercaptoethanol, N-ethylmaleimide, iodoacetic acid and iodoacetamide.

• Journal of Applied Biological Chemistry
• /
• 제42권3호
• /
• pp.107-112
• /
• 1999

A carboxymethyl-cellulase (CMCase) was purified from the culture supernatant of Bacillus sp. KD1014 by ultrafiltration, ammonium sulfate precipitation, and a series of chromatography on QAE-Sephadex A-50, hydroxylapatite and Sephadex G-75. The purified CMCase was a single protein of 32 kDa, showed an optimum activity at $60^{\circ}C$ and pH 6.0, and had a half-life of 23 min at $70^{\circ}C$. The enzyme activity was not influenced by metal ions such as $Mg^{2+},\;Fe^{3+},\;K^+,\;Zn^{2+}$, and $Cu^{2+}$ at a concentration of 1.0 mM, partially inhibited by $Mn^{2+}$ and $Ag^+$, and significantly inhibited by pentachlorophenol (PCP). The purified enzyme showed a 3.9-times higher activity on lichenan than on CMC, but hardly cleaved xylan, starch, avicel, laminarin, filter paper and levan. The results of activity staining of the purified enzyme separated by native and denaturing gel electrophoresis suggested that the CMCase might exist in dimeric, oligomeric or aggregated form as well as in monomeric form. The enzymatic cleavage products from cellotetraose indicated that the CMCase possessed transglycosylation activity.

• Journal of Applied Biological Chemistry
• /
• 제59권4호
• /
• pp.373-377
• /
• 2016

An extracellular metalloprotease, Btmp, was partially purified from the culture supernatant of Bacillus thuringiensis 29-126, isolated from animal feces collected in a zoological garden in Japan, by ultrafiltration, ammonium sulfate precipitation, and a set of chromatography on Sephadex G-75 and High-Q. The molecular mass of the protease was estimated to be 60 kDa by SDS-PAGE. The enzyme showed optimum activity at $50^{\circ}C$ and pH 6.0, and had a half-life of 14 min at $50^{\circ}C$. The enzyme activity was not influenced by $Na^+$, $K^+$, $As^+$, $Mg^{+2}$, $Ca^{2+}$, $Ba^{2+}$, and phenylmethylsulfonyl fluoride, but it was moderately inhibited by $Zn^{+2}$ at a concentration of 1.0 mM, while the activity was significantly inhibited to less than 50 % by $Cu^{2+}$, $Co^{2+}$, $Cd^{2+}$, and ethylenediaminetetraacetic acid. Interestingly, the enzyme was activated to 178 % by 1.0 mM of $Mn^{2+}$. From these results, it may be suggested that the protease is a novel extracellular manganeseactivated metalloprotease.

• 약학회지
• /
• 제35권3호
• /
• pp.222-230
• /
• 1991

Polyphenol oxidase (PPO) was purified from an extract of Perillae Folium by ammonium sulfate fractionation and sephadex G-150 gel filtration, which molecular weight estimated 65,000$\pm$1,000 in SDS-gel electrophoresis, and pI value was 4.8. The pH and temperature optima were 6.0 and $30^{\circ}C$ respectively. $K_{m}$ values of the PPO for various phenolics derived from Lineweaver-Burk plots were 4.0$\times$10$^{-4}$, caffeic acid; 4.2$\times$10$^{-3}$M, 4-methylcatechol. The inhibition by 4-nitrocatechoi, potassium cyanide, cysteine, 2-mercaptoethanol was competitive with $K_{i}$ values of 7.6$\times$10$^{-5}$M, 7.2$\times$10$^{-5}$M, 3.6$\times$10$^{-5}$M, 2.2$\times$10$^{-5}$M, respectively. Among the divalent cations, Cu$^{2+}$ ion was strong activator on PPO and Zn$^{2+}$, Ni$^{2+}$ ions were little effect on PPO activity. In comparing the amino acid composition of Perillae Folium PPO with that of wheat isozyme, grape, spinach showed similarity. But the content of glycine phenylalanine was abundant relatively.