• Applied Biological Chemistry
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• 제27권4호
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• pp.271-277
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• 1984

Pseudomonas stutzeri IAM 12097이 생산(生産)하는 Exo-maltotetraohydrolase의 gel filtration에 의하여 추정(推定)된 분자량(分子量)은 60,000이었고, SDS-polyacrylamide gel electrophoresis에 의하여 추정(推定)된 분자량(分子量)은 63,000이었으며 등전점(等電點)은 PH 4.8이었다. 최적(最適)pH는 6.6, PH안정(安定)범위는 $6.0{\sim}10.5$, 최적온도(最適溫度)는 $45{\sim}50^{\circ}C,\;40^{\circ}C$이하(以下)에서는 안정(安定)하였으며, $55^{\circ}C$이상(以上)에서는 급격하게 불활성화(不活性化)되었다. 본효소(本酵素)는 $Ag^+,\; Hg^{++},\;I_2,\;{\beta}-cycoldextrin$에 의하여 완전(完全)히 저해(沮害)되었고 EDTA, ${\rho}-CMB$, IAA에 의하여 약간 저해(沮害)되었다. soluble starch, amylopectin, amylose에 대(對)한 Michaelis constant(Km)는 각각(各各) 7.70mg/ml, 6.17mg/ml, 5.56mg/ml이었다.

• BMB Reports
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• 제29권5호
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• pp.455-461
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• 1996

A serine proteinase was purified from Acanthamoeba culbertsoni by 41~80% ammonium sulfate fractionation, ion exchange chromatography, affinity chromatography and gel filtration chromatography. The molecular weight of the purified enzyme was estimated to be 108.0 kDa by gel filtration chromatography and 54.0 kDa by SDS-PAGE. Therefore, the purified enzyme seemed to be a dimer. Isoelectric point was 4.5. The enzyme activity was highly inhibited by the serine proteinase inhibitors diisopropyl fluorophosphate (OFP) and phenylmethyl sulfonylfluoride (PMSF). It had a narrow pH optimum of 6.5~7.5 with a maximum at pH 7.0. These data suggested that the purified enzyme was a neutral serine proteinase. Optimal temperature was $37^{\circ}C$. It was stable for at least 16 h at $4^{\circ}C$ and $37^{\circ}C$, but it was rapidly inactivated at $65^{\circ}C$ The activity of the purified enzyme was not influenced significantly by $Mg^{2+}$, $Mn^{2+}$, $Zn^{2+}$ or $Ca^{2+}$. However, the enzyme activity was highly inhibited by $Hg^{2+}$ The enzyme degraded type I collagen and fibronectin, but not BSA, hemoglobin, lysozyme, immunoglobulin A or immunoglobulin G.

• BMB Reports
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• 제30권1호
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• pp.46-54
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• 1997

Alkaliphilic Bacillus sp. S-1 secretes a large amount (approximately 80% of total pullulanase activity) of an extracellular pullulanase (PUL-E). The pullulanase exists in two forms: a precursor form (PUL-I: $M_r$ 180,000), and a processed form (PUL-E: $M_r$ 140,000). Two forms were purified to homogeneity and their properties were compared. PUL-I was different in molecular weight, isoelectric point, $NH_2$-terminal amino acid sequence, and stabilities over pH and temperature ranges. The catalytic activities of PUL-I were also distinguishable in the $K_m$ and $V_{max}$ values for various substrates, and in the specific activity for pullulan hydrolysis. PUL-E showed 10-fold higher specific activities than PUL-I. However. PUL-I is immunologically identical to PUL-E, suggesting that PUL-I is initially synthesized and proteolytically processed to the mature form of PUL-E. Processing was inhibited by PMSF, but not by pepstatin, suggesting that some intracellular serine proteases could be responsible for processing of the PUL-I. PUL-I has a different conformational structure for antibody recognition from that of PUL-E. It is also postulated that the translocation of alkaline pullulanase(AP) in the bacterium possibly requires processing of the $NH_2$-terminal region of the AP protein. Processing of the precursor involves a conformational shift. resulting in a mature form. Therefore. precursor processing not only cleaves the signal peptide, but also induces conformational shift. allowing development of active form of the enzyme.

• BMB Reports
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• 제37권6호
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• pp.684-690
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• 2004

Heparin lyase I was purified to homogeneity from Bacteroides stercoris HJ-15 isolated from human intestine, by a combination of DEAE-Sepharose, gel-filtration, hydroxyapatite, and CM-Sephadex C-50 column chromatography. This enzyme preferred heparin to heparan sulfate, but was inactive at cleaving acharan sulfate. The apparent molecular mass of heparin lyase I was estimated as 48,000 daltons by SDS-PAGE and its isoelectric point was determined as 9.0 by IEF. The purified enzyme required 500 mM NaCl in the reaction mixture for maximal activity and the optimal activity was obtained at pH 7.0 and $50^{\circ}C$. It was rather stable within the range of 25 to $50^{\circ}C$ but lost activity rapidly above $50^{\circ}C$. The enzyme was activated by $Co^{2+}$ or EDTA and stabilized by dithiothreitol. The kinetic constants, $K_m$ and $V_{max}$ for heparin were $1.3{\times}10^{-5}\;M$ and $8.8\;{\mu}mol/min{\cdot}mg$. The purified heparin lyase I was an eliminase that acted best on porcine intestinal heparin, and to a lesser extent on porcine intestinal mucosa heparan sulfate. It was inactive in the cleavage of N-desulfated heparin and acharan sulfate. In conclusion, heparin lyase I from Bacteroides stercoris was specific to heparin rather than heparan sulfate and its biochemical properties showed a substrate specificity similar to that of Flavobacterial heparin lyase I.

• 공업화학
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• 제8권5호
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• pp.830-836
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• 1997

본 실험에서는 물유리를 사용하여 산세척에 의하여 제조된 산화철 입자의 표면개질에 대하여 연구하였다. 사용한 물유리의 $SiO_2$와 $Na_2O$의 몰비($SiO_2/Na_2O$)는 1, 2, 3.5이였다. 첨가되는 실리카의 양과 pH에 따라 산화철 현탁액의 분산성을 입자의 표면하전과 침강속도에 의하여 평가하였다. 그리고, 중성 영역에서 산화철 입자의 분산안정성을 유지할 수 있는 표면개질제(실리카)의 양을 도출하였으며, 물유리에 의한 산화철 입자의 표면개질을 습식 볼밀링에 의하여 슬러리 상태에서 실시하였다. 그 결과, 표면처리한 산화철 현탁액의 분산 안정성은 실리카의 양과 pH에 상호 의존하였다. 미처리한 산화철은 등전점인 pH 8에서 분산안정성을 잃고 있었으나, 산화철에 대하여 약 0.8wt%의 실리카로 표면처리한 산화철은 pH 5 이상 중성영역에서 분산안정성을 나타내었으며, 음이온성 계면활성제를 0.2wt% 이상 첨가에 의한 분산안정성이 더욱 증가되었다.

• Journal of Microbiology and Biotechnology
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• 제15권1호
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• pp.72-79
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• 2005

A new Bacillus peptidase, bacillopeptidase DJ-2 (bpDJ-2), with molecular mass of 42 kDa and isoelectric point (pI) of 3.5- 3.7, was purified to homogeneity from Bacillus sp. DJ-2 isolated from Doen-Jang, a traditional Korean soybean fermented food. The enzyme was identified as an extracellular serine fibrinolytic protease. The optimal conditions for the reaction were pH 9.0 and $60^{\circ}C$. The first 18 amino acid residues of the N-terminal amino acid sequence of bpDJ-2 were TDGVEWNVDQIDAPKAW, which is identical to that of bacillopeptidase F (bpf). However, based on their Nterminal amino acid sequence, molecular size, and pI, it is different from that of bpf and extracellular 90 kDa. The whole (2,541 bp, full-bpDJ-2) and mature (1,956 bp, mature-bpDJ-2) genes were cloned, and its nucleotide sequence and deduced amino acid sequence were determined. The expressed proteins, full-bpDJ-2 and mature-bpDJ-2, were detected on SDSPAGE at expected sizes of 92 and 68 kDa, respectively.

• 한국수산과학회지
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• 제31권3호
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• pp.393-398
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• 1998

멸치 액젓의 숙성 기간이 품질에 미치는 영향을 알아보기 위하여, 숙성 기간별에 따른 멸치의 총 질소 함량과 미분해성 펩티드의 분포와 특성 등을 semi-micro Kjeldahl 질소정량법, SDS-PAG 전기영동 및 등전점 전기영동법으로 알아보고, 이 중 전보에서 확인된 특정 펩티드를 품질의 지표로 설정할 때, 이에 미치는 숙성기간의 영향을 검토하였다. 아울러 특정 펩티드의 면적비가 물로 희석한 액젓의 희석 비율을 정확히 반영하는지의 여부도 검토하였다. 수분은 숙성 10개월 이후, $65\%$전후를 유지하였고, 총질소 함량은 숙성기간에 비례하여 증가하다가 18개월 이후는 증가 속도가 둔화하였다. 한편 단백질 함량은 숙성기간에 따라 감소하였고, 아미노산의 함량은 숙성 10개월 이후에는 변화가 거의 없었다. 분자량 55,600 dalton과 46,900 dalton의 펩티드는 숙성기간에 관계없이 나타나, 이들 펩티드가 멸치 액젓의 품질 지표로 활용될 수 있음을 보여주고 있었다. 한편 등전점 5.2, 5.6 및 6.0의 펩티드는 숙성기간에 관계없이 모든 시료에서 나타났다. 숙성기간에 따른 총질소 함량과 단백질 함량의 변화는 역의 상관관계를 나타내고 있었으며, 아미노산 함량과 SDS-PAG전기영동에 나타난 분자량 55,600 dalton의 펩티드 면적비와는 거의 상관성이 없는 것으로 나타났다 따라서 8-24개월 동안의 숙성 중에도 거의 영향을 받지않는 특정 펩티드의 면적비는 멸치 액젓의 품질 지표로 활용이 가능할 것으로 판단되었으며, 또한 액젓의 희석에 따른 총 질소 함량의 변화와 SDS-PAG 전기영동상의 펩티드 면적비의 변화는 희석비를 정확하게 반영하는 것으로 나타났다.

• Journal of Plant Biology
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• 제39권3호
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• pp.173-177
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• 1996

A novel calcium binding protein (CaBP) was purified to electrophoretic homogeneity from Dunaliella salina. In the course of purification experiment, this CaBP was identified as a monomer and its molecular weight was about 21 kDand isoelectric point (pI) value was about 4.1 using isoelectrofocusing. This CaBP was able to bind Ca2+ even in the pressence of an excess MgCl2 and KCI both in solution. In the SDS-PAGE, the Ca2+-bound form was slower than the Ca2+-free form in the nondenaturing PAGE. This means that the CaBP undergoes conformational change in the Ca2+-bound condition. Furthermore, UV absorption spectrum and fluorescence intensity of this CaBP was investigated. UV absorption peak was appeared at about 258 nm and decreased somewhat in Ca2+-bound condition. In the measurement of fluorescence, maximum intensity was appeared at 303 nm and decreased in Ca2+-bound state, similarly as UV absorption spectrum. These show distinct changes upon Ca2+-binding, which indicate of structural and/or dynamic changes largely reminiscent of other members of the EF-hand Ca2+-binding protein family.

• Membrane and Water Treatment
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• 제4권2호
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• pp.109-126
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• 2013

Surface modification by low-pressure ammonia ($NH_3$) plasma on commercial thin-film composite (TFC) membranes was investigated in this study. Surface hydrophilicity, total surface free energy, ion exchange capacity (IEC) and zeta (${\zeta}$)-potentials were determined for the TFC membranes. Qualitative and quantitative analyses of the membrane surface chemistry were conducted by attenuated total reflectance Fourier transform infrared (ATR FT-IR) spectroscopy. Results showed that the $NH_3$ plasma treatment increased the surface hydrophilicity, in particular at a plasma treatment time longer than 5 min at 50 W of plasma power. Total surface free energy was influenced by the basic polar components introduced by the $NH_3$ plasma, and isoelectric point (IEP) was shifted to higher pH region after the modification. A ten (10) min $NH_3$ plasma treatment at 90 W was found to be adequate for the TFC membrane modification, resulting in a membrane with better characteristics than the TFC membranes without the modification for water treatment. The thin-film chemistry (i.e., fully-aromatic and semi-aromatic nature in the interfacial polymerization) influenced the initial stage of plasma modification.

• Biotechnology and Bioprocess Engineering:BBE
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• 제11권2호
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• pp.134-139
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• 2006

Methanotrophs are microorganisms that possess the unique ability to utilize methane as their sole source of carbon and energy. A novel culture system, known as the compulsory circulation diffusion system, was developed for rapid growth of methanotrophic bacteria. Methanol dehydrogenase (MDH, EC 1.1.99.8) from Methylomicrobium sp. HG-1, which belongs to the type I group of methanotrophic bacteria, can catalyze the oxidation of methanol directly into formaldehyde. This enzyme was purified 8-fold to electrophoretic homogeneity by means of a 4 step procedure and was found in the soluble fraction. The relative molecular weight of the native enzyme was estimated by gel filtration to be 120 kDa. The enzyme consisted of two identical dimers which, in turn, consisted of large and small subunits in an ${\alpha}_2{\beta}_2$ conformation. The isoelectric point was 5.4. The enzymatic activity of purified MDH was optimum at pH 9.0 and $60^{\circ}C$, and remained stable at that temperature for 20 min. MDH was able to oxidize primary alcohols from methanol to octanol and formaldehyde.