• Title/Summary/Keyword: Molecular Characteristics

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Aspergillus niger가 생성하는 생전분 분해효소의 정제와 특성

  • 정만재
    • Microbiology and Biotechnology Letters
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    • v.25 no.2
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    • pp.166-172
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    • 1997
  • Aspergillus niger was selected as a strain producing the potent raw starch hydorlyzing enzyme. These experiments were conducted to investigate the conditions of the glucoa- mylase production, the purification of the enzyme, some characteristics of the purified enzyme and hydrolysis rate on various raw starches such as com, rice, potato, glutinous rice, sweet potato, wheat and barley. The optimum cultural temperature and time for the enzyme production on wheat bran medium were $30^{\circ}C$ and 96hrs, respectively. The respective addition of yeast extract and nutrient broth on wheat bran medium increased slightly the enzyme production. The enzyme was purified by ammonium sulfate fractionation and DEAE-cellulose column chromatography. The specific activity of the purified enzyme was 30.7u/mg-protein and the yield of enzyme activity was 25.8%. The purified enzyme showed a single band on polyacrylamide disc gel electrophoresis and its molecular weight was estimated to be 56,000 by SDS-polyacrylamide disc gel electrophoresis. The isoelectric point for the purified enzyme was pH3.7. The optimum temperature and pH were $65^{\circ}C$ and pH 4.0, respectively. The purified enzyme was stable in the pH range of pH 3.0-9.5 and below $45^{\circ}C$, and its thermal stability was slightly increased by the addition of $Ca^{2+}$. The purified enzyme was activated by $Co^{2+},\;Sr^{2+},\;Mn^{2+},\;Fe^{2+},\;Cu^{2+}$. Raw rice starch, raw corn starch, raw glutinous rice starch, raw sweet potato starch, raw wheat starch and raw barley starch showed more than 90% hydrolysis rate in 48hrs incubation. Even raw potato starch, most difficult to be hydrolyzed, showed 80% hydrolysis rate. The purified enzyme was identified as glucoamylase.

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Characteristics of Raw Starch-Digesting Enzyme from Streptomyces sp. 4M-2 (Streptomyces sp. 4M-2가 생산하는 생전분 분해효소의 특성)

  • 최성현;김찬조;오만진;이종수
    • Microbiology and Biotechnology Letters
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    • v.17 no.2
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    • pp.136-141
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    • 1989
  • A raw starch-digesting enzyme from Streptomyces sp. 4M-2 was purified by ammonium sulfate fractionation, DEAE-Sephadex A-50 column chromatography and Sephadex G-100 gel filtration. The specific activity of the purified enzyme was 51.22 RSU/mg protein and the yield was 4.5% of the total activity of the culture broth. The purified enzyme was found to be homogeneous by polyacrylamide gel electrophoresis and its molecular weight was estimated to be about 102, 000 daltons by SDS-polyacrylamide gel electrophoresis, The optimal temperature and pH for the enzyme activity were 42$^{\circ}C$ and PH 5.5, respectively. The enzyme had Km, value of 44.44mg/$m\ell$ for raw corn starch. The enzyme was activated by addition of calcium and barium ions. Corn amylose was degraded by the enzyme very easily and raw potato starch was also degraded easily. Main products of the enzymatic hydrolysis of raw corn starch were analyzed to be maltose and maltotriose. The enzyme was considered as $\alpha$-amylase.

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Heterologous Expression and Optimized One-Step Separation of Levansucrase via Elastin-like Polypeptides Tagging System

  • Kang, Hye-Jin;Kim, Jin-Hee;Chang, Woo-Jin;Kim, Eung-Soo;Koo, Yoon-Mo
    • Journal of Microbiology and Biotechnology
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    • v.17 no.11
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    • pp.1751-1757
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    • 2007
  • Elastin-like polypeptides (ELPs) undergo a reversible inverse phase transition upon a change in temperature. This thermally triggered phase transition allows for a simple and rapid means of purifying a fusion protein. Recovery of ELPs-tagged fusion protein was easily achieved by aggregation, triggered either by raising temperature or by adding salt. In this study, levansucrase has been used as a model enzyme in the development of a simple one-step purification method using ELPs. The levansucrase gene cloned from Pseudomonas aurantiaca S-4380 was tagged with various sizes of ELPs to functionally express and optimize the purification of levansucrase. One of two ELPs, ELP[V-20] or ELP[V-40], was fused at the C-terminus of the levansucrase gene. A levansucrase-ELP fusion protein was expressed in Escherichia coli $DH5{\alpha}$ at $37^{\circ}C$ for 18 h. The molecular masses of levansucrase-ELP[V-20] and levansucrase-ELP[V-40] were determined as 56 kDa and 65 kDa, respectively. The phase transition of levansucrase-ELP[V-20] occurred at $20^{\circ}C$ in 50 mM Tris-Cl (pH 8) buffer with 3 M NaCl added, whereas the phase transition temperature ($T_t$) of levansucrase-ELP[V-40] was $17^{\circ}C$ with 2 M NaCl. Levansucrase was successfully purified using the phase transition characteristics of ELPs, with a recovery yield of higher than 80%, as verified by SDS-PAGE. The specific activity was measured spectrophotometrically to be 173 U/mg and 171 U/mg for levansucrase-ELP[V-20] and levansucrase-ELP[V-40], respectively, implying that the ELP-tagging system provides an efficient one-step separation method for protein purification.

Variation of Natural Organic Matter Characteristics through Water Treatment Processes (정수공정별 천연유기물질의 특성 변화)

  • Hwang, Jeong-Eun;Kang, Lim-Seok;Kim, Seung-Hyun;Yoon, Cho-Hee
    • Journal of Korean Society of Environmental Engineers
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    • v.22 no.7
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    • pp.1253-1261
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    • 2000
  • Natural organic matter (NOM) which occurs ubiquitously in both surface and ground waters, consists of both humic (i.e., humic and fulvic acids) and nonhumic components. NOM in general as well as certain constituents are problematic in water treatment. From a regulatory perspective, concerns focus upon the role of NOM constituents as disinfection byproduct (DBP) precursors. The fractionation of NOM through water treatment processes can provide insight into treatment process selection and applicability. Problematic NOM fractions can be targeted for removal or transformation. Significant source-related differences in NOM were observed among various source waters. This study found that bulk Dissolved Organic Carbon (DOC) concentration was hardly removed by oxidation process. Oxidation transformed high Molecular Weight (MW) hydrophobic fraction into low MW hydrophilic fraction. Ozone reduced s-pecific Ultraviolet Absorbance (SUVA) value more than chlorine. High MW hydrophobic fraction was effectively removed by coagulation process. About 50% of Trihalomethane Formation Potential (THMFP) was removed by coagulation process.

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Hydroxylation of Compactin (ML-236B) by CYP105D7 (SAV_7469) from Streptomyces avermitilis

  • Yao, Qiuping;Ma, Li;Liu, Ling;Ikeda, Haruo;Fushinobu, Shinya;Li, Shengying;Xu, Lian-Hua
    • Journal of Microbiology and Biotechnology
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    • v.27 no.5
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    • pp.956-964
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    • 2017
  • Compactin and pravastatin are competitive cholesterol biosynthesis inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase and belong to the statin drugs; however, the latter shows superior pharmacokinetic characteristics. Previously, we reported that the bacterial P450, CYP105D7, from Streptomyces avermitilis can catalyze the hydroxylation of 1-deoxypentalenic acid, diclofenac, and naringenin. Here, we demonstrate that CYP105D7 could also catalyze compactin hydroxylation in vitro. In the presence of both bacterial and cyanobacterial redox partner systems with an NADPH regeneration system, the reaction produced two hydroxylated products, including pravastatin (hydroxylated at the C6 position). The steady-state kinetic parameters were measured using the redox partners of putidaredoxin and its reductase. The $k_m$ and $k_{cat}$ values for compactin were $39.1{\pm}8.8{\mu}M$ and $1.12{\pm}0.09min^{-1}$, respectively. The $k_{cat}/K_m$ value for compactin ($0.029min^{-1}{\cdot}{\mu}M^{-1}$) was lower than that for diclofenac ($0.114min^{-1}{\cdot}{\mu}M^{-1}$). Spectroscopic analysis showed that CYP105D7 binds to compactin with a $K_d$ value of $17.5{\pm}3.6{\mu}M$. Molecular docking analysis was performed to build a possible binding model of compactin. Comparisons of different substrates with CYP105D7 were conclusively illustrated for the first time.

Detecting of Periodic Fasciculations of Avian Muscles Using Magnetic and Other Multimedia Devices

  • Nakajima, Isao;Tanaka, Sachie;Mitsuhashi, Kokuryo;Hata, Jun-ichi;Nakajima, Tomo
    • Journal of Multimedia Information System
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    • v.6 no.4
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    • pp.293-302
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    • 2019
  • In the past, there was a theory that influenza wasn't transmitted directly from birds but was infected to humans via swains. Recently, molecular level research has progressed, and it was confirmed that the avian influenza virus can directly infected to human lung and intestinal epithelial cells. Three pandemicsin the past 100 years were also infected to humans directly from birds. In view of such scientific background, we are developing a method for screening sick birds by monitoring the physiological characteristics of birds in a contactless manner with sensors. Here, the movement of respiratory muscles and abdominal muscles under autonomic innervation was monitored using a magnet and Hall sensor sewn on the thoracic wall, and other multimedia devices. This paper presents and discusses the results of experiments involving continuous periodic noise discovered during flight experiments with a data logger mounted on a Japanese pheasant from 2012 to 2015. A brief summary is given as the below: 1. Magnet and Hall sensor sewn to the left and right chest walls, bipolar electrocardiograms between the thoracic walls, posterior thoracic air sac pressure, angular velocity sensors sewn on the back and hips, and optical reflection of LEDs (blue and green) from the skin of the hips allow observation of periodic vibrations(fasciculations) in the waves. No such analysis has been reported before. 2. These fasciculations are presumed to be derived from muscle to maintain and control air sac pressure. 3. Since each muscle fiber is spatially Gaussian distributed from the sympathetic nerve, the envelope is assumed to plot a Gaussian curve. 4. Since avian trunk muscles contract periodically at all time, we assume that the sympathetic nerve dominates in their control. 5. The technique of sewing a magnet to the thoracic wall and measuring the strength of the magnetic field with a Hall sensor can be applied to screen for early stage of avian influenza, with a sensor attached to the chicken enclosure.

Penicillium menonorum: A Novel Fungus to Promote Growth and Nutrient Management in Cucumber Plants

  • Babu, Anam Giridhar;Kim, Sang Woo;Yadav, Dil Raj;Hyum, Umyong;Adhikari, Mahesh;Lee, Youn Su
    • Mycobiology
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    • v.43 no.1
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    • pp.49-56
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    • 2015
  • The present study is the first report on the isolation of Penicillium menonorum from rhizosphere soil in Korea and its identification based on morphological characteristics and internal transcribed spacer gene sequence. The fungal isolate was named KNU-3 and was found to exhibit plant growth-promoting (PGP) activity through indole acetic acid (IAA) and siderophore production, as well as P solubilization. KNU-3 produced 9.7 mg/L IAA and solubilized 408 mg of $Ca_3PO_4/L$, and inoculation with the isolate significantly (p < 0.05) increased the dry biomass of cucumber roots (57%) and shoots (52%). Chlorophyll, starch, protein, and P contents were increased by 16%, 45%, 22%, and 14%, respectively, compared to plants grown in uninoculated soil. The fungus also increased soil dehydrogenase (30%) and acid phosphatase (19%) activities. These results demonstrate that the isolate KNU-3 has potential PGP attributes, and therefore it can be considered as a new fungus to enhance soil fertility and promote plant growth. Moreover, the discovery of PGP ability and traits of this fungus will open new aspects of research and investigations. In this study, plant growth promotion by P. menonorum KNU-3 is reported for the first time in Korea after its original description.

Sclerotium Rot of Cowpea (Vigna sinensis King) Caused by Sclerotium rolfsii (Sclerotium rolfsii에 의한 동부 흰비단병)

  • Kwon, Jin-Hyeuk;Kang, Dong-Wan;Han, Inyoung;Choi, Yong-Jo;Lee, Sang-Dae;Son, Daeyoung
    • The Korean Journal of Mycology
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    • v.44 no.1
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    • pp.61-63
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    • 2016
  • Sclerotium rot disease on cowpea (Vigna sinensis King) was observed in the exhibition field of Gyeongsangnam-do Agricultural Research and Extension Services in September 2015. Lesions were covered by white mycelial mats, and numerous sclerotia were formed on the stem near the soil line. The sclerotia were globoid in shape, 1~3 mm in size and white to brown in color. The optimum temperature for mycelial growth and sclerotia formation on potato dextrose agar (PDA) was $30^{\circ}C$, with the hyphal width of $4{\sim}8{\mu}m$. For molecular identification, the complete internal transcribed spacer (ITS) rDNA region of the causal fungus was sequenced and analyzed. Based on the mycological characteristics, ITS rDNA sequence analysis, and pathogenicity test, this fungus was identified as Sclerotium rolfsii. This is the first report of sclerotium rot on cowpea caused by S. rolfsii in Korea.

Unrecorded Fungal Species Isolated from Greenhouses Used for Shiitake Cultivation in Korea (표고버섯 재배사에서 분리한 국내 미기록 진균 보고)

  • Ahn, Geum Ran;Kwon, Hyuk Woo;Ko, Han Kyu;Kim, Seong Hwan
    • The Korean Journal of Mycology
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    • v.44 no.1
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    • pp.8-15
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    • 2016
  • Fungal contamination is a detrimental factor affecting sawdust media-based shiitake cultivation in greenhouses. During fungal monitoring of greenhouses used for shiitake cultivation, eight fungal species were isolated and identified from indoor air and mushroom flies collected in the greenhouses. The current study reported five species as new in Korea, viz. Ascochyta hordei, Discosia artocreas, Mucor nidicola, Perenniporia medulla-panis, and Pseudozyma prolifica, and confirmed two species, Penicillium charlesii and Penicillium brevicompactum, which were previously recorded in Korea without molecular taxonomic validation. The morphological characteristics and phylogenetic relationships based on nucleotide sequences of the internal transcribed spacer rDNA region or calmodulin gene were described for all identified species.

A Report of Eighteen Unrecorded Fungal Species in Korea (국내에서 분리된 미기록 진균 18종 보고)

  • Ahn, Geum Ran;Choi, Min Ah;Kim, Ji Eun;Seo, Eun Ji;Kim, Jun Young;Kim, Seong Hwan
    • The Korean Journal of Mycology
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    • v.45 no.4
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    • pp.292-303
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    • 2017
  • During a survey of culturable fungi in Korea, 18 unrecorded fungal species were isolated and identified from the indoor air of mushroom cultivation houses, the materials used for preparation of mushroom cultivation media, wild plants, and funitures. This study reports the descriptions of the 18 unrecorded fungal species: Aspergillus creber, Ceratocystis paradoxa, Colletotrichum spaethianum, Coniochaeta velutina, Coprinellus xanthothrix, Epicoccum sorghinum, Leptosphaeria rubefaciens, Myrothecium gramineum, Paraconiothyrium fuckelii, Penicillium erubescens, Penicillium melinii, Penicillium pulvillorum, Penicillium sabulosum, Penicillium turbatum, Pestalotiopsis portugalica, Pilidiella castaneicola, Rachicladosporium pini, and Umbelopsis nana. For all the identified species, the morphological characteristics including the features of colony formed on media, images of light microscopy, and molecular phylogenetic relationships based on nucleotide sequences of the internal transcribed spacer ribosomal DNA (ITS rDNA), 18S rDNA, 28S rDNA, ${\beta}-tubulin$ gene, calmodulin gene, and translation elongation factor gene were described.