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Genenation of structural diversity in polyketides by combinatorial biosynthesis of polyketides: Part I. Generation of multiple bioactive macrolides by hybrid modular polyketide synthases in Streptomyces venezuelae, Part II. Production of novel rifamycins by combinatorial biosynthesis

  • Yoon, Yeo-Joon
    • Proceedings of the Korean Society for Applied Microbiology Conference
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    • 2002.10a
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    • pp.18-25
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    • 2002
  • The pikromycin biosynthetic system in Streptomyces venezuleae is unique for its ability to produce two groups of antibiotics that include the 12-membered ring macrolides methymycin and neomethymycin, and the 14-membered ring macrolides narbomycin and pikromycin. The metabolic pathway also contains two post polyketide-modification enzymes, a glycosyltransferase and P450 hydroxylase that have unusually broad substrate specificities. In order to explore further the substrate flexibility of these enzymes a series of hybrid polyketide synthases were constructed and their metabolic products characterized. The plasmid-based replacement of the multifunctional protein subunits of the pikromycin PKS in S. venezuelae by the corresponding subunits from heterologous modular PKSs resulted in recombinant strains that produce both 12- and 14-membered ring macrolactones with predicted structural alterations. In all cases, novel macrolactones were produced and further modified by the DesVII glycosyltransferase and PikC hydroxylase leading to biologically active macrolide structures. These results demonstrate that hybrid PKSs in S. venezuelae can produce a multiplicity of new macrolactones that are modified further by the highly flexible DesVII glycosyltransferase and PikC hydroxylase tailoring enzymes. This work demonstrates the unique capacity of the S. venezuelae pikromycin pathway to expand the toolbox of combinatorial biosynthesis and to accelerate the creation of novel biologically active natural products. The polyketide backbone of rifamycin B is assembled through successive condensation and ${\beta}$-carbonyl processing of the extender units by the modular rifamycin PKS. The eighth module, in the RifD protein, contains nonfunctional DH domain and functional KR domain, which specify the reduction of the ${\beta}$-carbonyl group resulting in the C-21 bydroxyl of rifamycin B. A four amino acid substitution and one amino acid deletion were introduced in the putative NADPH binding motif in the proposed KR domain encoded by rifD. This strategy of mutation was based on the amino acid sequences of the corresponding motif of the KR domain of module 3 in the RifA protein, which is believed dysfunctional, so as to introduce a minimum alteration and retain the reading frame intact, yet ensure loss of function. The resulting strain produces linear polyketides, from tetraketide to octaketide, which are also produced by a rifD disrupted mutant as a consequence of premature termination of polyketide assembly. Much of the structural diversity within the polyketide superfamily of natural products is due to the ability of PKSs to vary the reduction level of every other alternate carbon atom in the backbone. Thus, the ability to introduce heterologous reductive segments such as ketoreductase (KR), dehydratase (DH), and enoylreductase (ER) into modules that naturally lack these activities would increase the power of the combinatorial biosynthetic toolbox. The dehydratase domain of module 7 of the rifamycin PKS, which is predicted to be nonfunctional in view of the sequence of the apparent active site, was replaced with its functional homolog from module 7 of rapamycin-producing polyketide synthase. The resulting mutant strain behaved like a rifC disrupted mutant, i.e., it accumulated the heptaketide intermediate and its precursors. This result points out a major difficulty we have encountered with all the Amycolatopsis mediterranei strain containing hybrid polyketide synthases: all the engineered strains prepared so far accumulate a plethora of products derived from the polyketide chain assembly intermediates as major products instead of just analogs of rifamycin B or its ansamycin precursors.

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An Experimental Study on the Performance of Compression-Type Anchor for CFRP Tendons (CFRP 긴장재용 압착형 정착장치의 정착성능에 관한 실험적 연구)

  • Jung, Woo-Tai;Lee, Seung-Joo;Park, Young-Hwan
    • Journal of the Korea Concrete Institute
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    • v.20 no.5
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    • pp.611-618
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    • 2008
  • CFRP (carbon fiber reinforced polymer) tendons can be used as an alternative to solve the corrosion problem of steel tendons. Since CFRP tendons are vulnerable to transverse pressure and stress concentration, the conventional anchorage system used for steel tendons can create an unreliable load carrying capacity and may result in a premature failure. Therefore, it is necessary to develop the anchorage system that is well suited for CFRP tendons. There are many types of anchorage systems for CFRP tendons, which can be classified into three types: wedge-type anchorage, bond-type anchorage, and compression-type anchorage. This paper deals with the compression-type anchorage system manufactured through swaging technology. Based on the previous test results performed by the authors, the dimension of anchorage sleeve, the use and non-use of the insert, and the compression pressure on the sleeve have been selected as the major parameters affecting the performance of the compression-type anchorage. Some anchorage sleeves have been tapered to reduce the stress concentration. Test results revealed that the performance of the anchorage system depends mainly on the dimension and the compression pressure. It has been verified that the tapered sleeve can effectively reduce the stress concentration.

A prescription study in 『Dongeuibogam』 for the Applications of Cnidii Rhizoma (천궁(川芎)의 활용(活用)을 위한 『동의보감(東醫寶鑑)』 처방연구(處方硏究))

  • Jang, A-Ryoeng;Lee, Jin-Ho;Kim, Tae-Hyun;Kim, Dong-Hyun;Choi, Hyung-Wook;Jung, Myung;Yun, Yong-Gab;Lim, Kyu-Sang
    • Herbal Formula Science
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    • v.22 no.1
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    • pp.13-32
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    • 2014
  • Objectives : Until now the study of Cnidii Rhizoma, hemorrhage, brain waves, such as ischemic brain injury, analgesic, effect overcome of the stress from pregnancy melanin formation and inhibiting effects skin whitening have been published regarding this article. Cnidii Rhizoma demonstrates its different abilities depending on the characteristics. This paper reported that effect of Cnidii Rhizoma in Dongeuibogam blended prescriptions as main medicine. In addition, by analyzing data, we studied about utilizing of Cnidii Rhizoma. Methods : Cnidii Rhizoma in Dongeuibogam Prescriptions as the main ingredient was built with database of 202 prescriptions. Thus analyzed data was summarized in detail.(Table-1) If there is no difference in the title of the prescription but in other case the configuration information is different, formulations 1 and 2 were divided by the table. Results : The following results were reached through investigations on the prescriptions usikng Cnidii Rhizoma as a key component. 1. Prescriptions taking Cnidii Rhizoma as a monarch drug are utilized for 40 therapeutic purposes. In particular, 12.3% of prescriptions appear in the chapter of head, and 10.8% of those appear in the chapter of women, and 9.4% of eye, 8.9% of child, 6.4% of wind disease respectively. 2. Prescriptions utilizing Cnidii Rhizoma as the main ingredient are used in the treatment of headache, dizziness and pregnancy hemorrhage fetal movement, premature birth and they are also used for treating 131 different types of disease. 3. The dosage of Cnidii Rhizoma in formulas is from 2pun(about 0.75g) to 5don(nearly 18.75g), however 1don(nearly 3.75g) has been taken the most for clinical application. 4. We find out that according to herbs or prescriptions, Cnidii Rhizoma has a variety of functions such as ascending & descending of energy. Samultang is the most useful base prescription which used the Cnidii Rhizoma as the main component. Conclusion : These results suggest that, Cnidii Rhizoma once-amount use (don nearly 3.75g) 4g in head, gynecology, ophthalmology, pediatrics and paralysis disease associated with oriental medicine resource development can be considered to be widely used These results suggest that Cnidii Rhizoma was used most with 1 don(4g) and can be widely used for the resource development to the disease such as brain, gynecology, ophthalmologhy, pediatrics and wind-associated symptoms.

A Single Nucleotide Deletion resulting in Frameshift in Two Korean Neonates with Thyroxine-Binding Globulin Deficiency (단일 뉴클레오타이드 결손으로 인한 Frameshift 돌연변이로 규명된 티록신결합글로불린 결핍증 1례)

  • Park, Sang-Joon;Suh, Jin-Soon;Jung, Min-Ho;Lee, Hee-Jin;Suh, Byung-Kyu;Lee, Won-Bae;Lee, Byung-Churl
    • Clinical and Experimental Pediatrics
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    • v.48 no.11
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    • pp.1252-1255
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    • 2005
  • Abnormalities in the levels of thyroxine-binding globulin (TBG) are not associated with clinical disease and they do not require treatment. Congenital TBG deficiency is inherited in an X-linked manner. To date, some complete and partial TBG variants and one polymorphism have been identified by analysis of the TBG gene. Two male neonates were referred to us because of their low $T_4$ levels that were noted on the neonatal screening test. They showed normal levels of free $T_4$ and TSH. Their serum TBG was not detectable and those values of their parents were within the normal ranges. The genomic DNA was extracted from their white blood cells and the four coding exons of the TBG gene were amplified by using polymerase chain reaction. Sequencing of the four coding regions and all the intron/exon junctions revealed a single nucleotide deletion of the first base of the codon 352 of the mature protein in both of the neonates. This mutation resulted in a frameshift and a premature stop codon (TGA) 374. Their mothers were shown to be heterozygotes. We detected a single nucleotide deletion resulting in a frameshift in two male Korean neonates who had complete TBG deficiency.

The clinical manifestations, the short- and long-term outcomes of Bartter syndrome (Bartter 증후군의 임상 양상과 장단기 치료 결과)

  • Park, Hye Won;Lee, Joo Hoon;Park, Young Seo
    • Clinical and Experimental Pediatrics
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    • v.50 no.12
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    • pp.1231-1240
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    • 2007
  • Purpose : Bartter syndrome is a renal tubular defect in electrolyte transport characterized by hypokalemia, metabolic alkalosis and other clinical signs and symptoms. The aims of this study were to analyze the clinical manifestations and the short- and long-term outcomes of Bartter syndrome. Methods : We retrospectively reviewed clinical history, laboratory finding of blood and urine, renal ultrasonography, and hearing tests of five patients who were diagnosed and managed with Bartter syndrome at Asan Medical Center from April 1992 to May 2007. We also evaluated height and body weight periodically after institution of therapy. Results : All patients had poor oral intake, failure to thrive and polyuria. Three of them had maternal history of polyhydramnios and premature delivery. The mean age at diagnosis was 11.8 months. All children presented with hypokalemia, metabolic alkalosis, hyperreninemia. Their blood pressures were normal. One patient had nephrocalcinosis on renal ultrasonography and all of them had normal result in hearing tests. After treatment with indomethacin or other prostaglandin inhibitors and potassium supplementation,their clinical features improved with catch-up growth and improvement in the development during long-term follow-up. Conclusion : We emphasize that early diagnosis and proper treatment in patient with Bartter syndrome are related to better prognosis.

Association of Dopamine D4 Receptor(DRD4) Gene Polymorphism with Korean Schizophrenic Patients (정신분열병에서 도파민 D4 수용체(DRD4) 유전자의 다형성)

  • Lee, Hong Seock;Lee, Min Soo;Han, Deock-Jeong;Lee, Heon-Jeong
    • Korean Journal of Biological Psychiatry
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    • v.7 no.2
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    • pp.152-158
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    • 2000
  • Background : No association between schizophrenia and dopamine D4 receptor polymorphisms have been reported. Despite these results, it is premature to exclude the association. It has been suggested that the susceptibility to develop schizophrenia could result from variation at a number loci which may interact or coact with each other. Therefore, we investigated a possible association of combinations of exon III 48bp polymorphism[D4E3] and exon I 12bp polymorphism of the DRD4 gene [D4E1] with schizophrenia. Methods : 207 unrelated Korean schizophrenic patients and 191 healthy controls were recruited. DRD4 genotype was established using the polymerase chain reaction. Statistical analysis consisted of ${\chi}^2$ tests for Hardy-Weinberg proportions and genotypic and allelic frequencies in the patients and control groups. Results : There were no statistically significant differences in the each polymorphisms between schizophrenics and controls. And all genotype frequencies were within Hardy-Weinberg expectations. When the combinations of the polymorphism in schizophrenia and controls were compared, however, there were significant differences at $A1A2^*2/4$ in the distributions of the combinations of D4E1 and D4E3(p<0.01). Conclusions : These findings suggest that the certain combination of D4E1 and D4E3($A1A2^*2/4$) has the protective role to a susceptibility for schizophrenia.

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Improvement of Anti-Corrosion Characteristics for Light Metal in Surface Modification with Sulfuric Acid Solution Condition (경금속 표면개질 시 황산 수용액 조건에 따른 내식성 개선 효과)

  • Lee, Seung-Jun;Han, Min-Su;Kim, Seong-Jong
    • Journal of Advanced Marine Engineering and Technology
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    • v.39 no.3
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    • pp.223-229
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    • 2015
  • Surface modification is a technology to form a new surface layer and overcome the intrinsic properties of the base material by applying thermal energy or stress onto the surface of the material. The purpose of this technique is to achieve anti-corrosion, beautiful appearance, wear resistance, insulation and conductance for base materials. Surface modification techniques may include plating, chemical conversion treatment, painting, lining and surface hardening. Among which, a surface modification process using electrolytes has been investigated for a long time in connection with research on its industrial application. The technology is highly favoured by various fields because it provides not only high productivity and cost reduction opportunities, but also application availability for components with complex geometry. In this study, an electrochemical experiment was performed on the surface of 5083-O Al alloy to determine an optimal electrolyte temperature, which produces surface with excellent corrosion resistance under marine environment than the initial surface. The experiment result, the modified surface presented a significantly lower corrosion current density with increasing electrolyte temperature, except for $5^{\circ}C$ of electrolyte temperature at which premature pores was created.

Development of a Short-term Failure Assessment of High Density Polyethylene Pipe Welds - Application of the Limit Load Analysis - (고밀도 폴리에틸렌 융착부에 대한 단기간 파손 평가법 개발 - 한계하중 적용 -)

  • Ryu, Ho-Wan;Han, Jae-Jun;Kim, Yun-Jae;Kim, Jong-Sung;Kim, Jeong-Hyeon;Jang, Chang-Heui
    • Transactions of the Korean Society of Mechanical Engineers A
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    • v.39 no.4
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    • pp.405-413
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    • 2015
  • In the US, the number of cases of subterranean water contamination from tritium leaking through a damaged buried nuclear power plant pipe continues to increase, and the degradation of the buried metal piping is emerging as a major issue. A pipe blocked from corrosion and/or degradation can lead to loss of cooling capacity in safety-related piping resulting in critical issues related to the safety and integrity of nuclear power plant operation. The ASME Boiler and Pressure Vessel Codes Committee (BPVC) has recently approved Code Case N-755 that describes the requirements for the use of polyethylene (PE) pipe for the construction of Section III, Division 1 Class 3 buried piping systems for service water applications in nuclear power plants. This paper contains tensile and slow crack growth (SCG) test results for high-density polyethylene (HDPE) pipe welds under the environmental conditions of a nuclear power plant. Based on these tests, the fracture surface of the PENT specimen was analyzed, and the fracture mechanisms of each fracture area were determined. Finally, by using 3D finite element analysis, limit loads of HDPE related to premature failure were verified.

Risk Factors and Nutritional Profiles Associated with Stunting in Children

  • Febriani, Andi Dwi Bahagia;Daud, Dasril;Rauf, Syarifuddin;Nawing, Herry Demokrasi;Ganda, Idham Jaya;Salekede, Setia Budi;Angriani, Hadia;Maddeppungeng, Martira;Juliaty, Aidah;Alasiry, Ema;Artaty, Ratna Dewi;Lawang, St Aizah;Ridha, Nadirah Rasyid;Laompo, Amiruddin;Rahimi, Rahmawaty;Aras, Jusli;Sarmila, Besse
    • Pediatric Gastroenterology, Hepatology & Nutrition
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    • v.23 no.5
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    • pp.457-463
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    • 2020
  • Purpose: To analyze risk factors and various nutrients associated with stunting among children aged 6-60 months. Methods: This is a case-control and cross-sectional study between 40 stunting cases and 40 controls. Data on possible risk factors associated with stunting were obtained through direct interviews and using a questionnaire. Examination of vitamin D, zinc, albumin, and ferritin levels was performed on both groups. Data were analyzed using IBM SPSS Statistics for Windows, Version 23.0 (IBM Co., Armonk, NY, USA) to determine risk factors for stunting and to assess the relationship between nutritional levels and stunting. Results: The incidence of stunting was highest in children aged 12-36 months. Children with low weight and very low weight for age comprised of 55% and 22.5%, respectively, of the study participants. The highest mother's educational level was junior high school (40%). History of low birth weight (LBW) was more commonly observed in the stunting group than that in the control group (25.0% and 7.5%, respectively; p=0.034, odds ratio, 0.310 [95% confidence interval, 0.122-0.789]). Approximately 7.5% of cases had premature birth. Exclusive breast feeding was found to be not correlated with stunting. The mean zinc level in the stunting group was 34.17 ng/mL, which was different from that in the control group (50.83 ng/mL) (p=0.023). Blood ferritin, vitamin D, albumin, and calcium levels were not strongly correlated with stunting. Conclusion: LBW is the main risk factor contributing to stunting and is strongly associated with low zinc level.

Effects of 7-MEGATM 500 on Oxidative Stress, Inflammation, and Skin Regeneration in H2O2-Treated Skin Cells

  • Song, In-Bong;Gu, Hyejung;Han, Hye-Ju;Lee, Na-Young;Cha, Ji-Yun;Son, Yeon-Kyong;Kwon, Jungkee
    • Toxicological Research
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    • v.34 no.2
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    • pp.103-110
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    • 2018
  • Environmental stimuli can lead to the excessive accumulation of reactive oxygen species (ROS), which is one of the risk factors for premature skin aging. Here, we investigated the protective effects of $7-MEGA^{TM}$ 500 (50% palmitoleic acid, 7-MEGA) against oxidative stress-induced cellular damage and its underlying therapeutic mechanisms in the HaCaT human skin keratinocyte cell line (HaCaT cells). Our results showed that treatment with 7-MEGA prior to hydrogen peroxide ($H_2O_2$)-induced damage significantly increased the viability of HaCaT cells. 7-MEGA effectively attenuated generation of $H_2O_2$-induced reactive oxygen species (ROS), and inhibited $H_2O_2$-induced inflammatory factors, such as prostaglandin $E_2$ ($PGE_2$), tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$), and $interleukin-1{\beta}$ ($IL-1{\beta}$). In addition, cells treated with 7-MEGA exhibited significantly decreased expression of matrix metalloproteinase-1 (MMP-1) and increased expression of procollagen type 1 (PCOL1) and Elastin against oxidative stress by $H_2O_2$. Interestingly, these protective activities of 7-MEGA were similar in scope and of a higher magnitude than those seen with 98.5% palmitoleic acid (PA) obtained from Sigma when given at the same concentration (100 nL/mL). According to our data, 7-MEGA is able to protect HaCaT cells from $H_2O_2$-induced damage through inhibiting cellular oxidative stress and inflammation. Moreover, 7-MEGA may affect skin elasticity maintenance and improve skin wrinkles. These findings indicate that 7-MEGA may be useful as a food supplement for skin health.