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Biochemical Characterizations of Phenylalanine Ammonia-Lyase and its Mutants to Develop an Enzymatic Therapy for Phenylketonuria (페닐케톤뇨증의 효소치료 개발을 위한 phenylalanine ammonia-lyase 및 유전자 변이형의 생화학적 특성)

  • Kim, Woo-Mi
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1226-1231
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    • 2009
  • Enzyme substitution with recombinant phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is currently being explored for treatment of phenylketonuria (PKU), an autosomal recessive genetic disorder with mutations of the gene encoding phenylalanine-4-hydroxylase (EC 1.14.16.1). However, oral administration of PAL is limited because of proteolytic digestion in the gastrointestinal tract. The aim of this study was to determine the biochemical properties of PAL and delinate the susceptibility of wild-type PAL to pancreatic proteolysis by exploring several mutants, and to develop therapeutic drugs with PAL for PKU. The specific activity of PAL was assayed and its optimal pH, temperature stability, and intestinal protease susceptibility were investigated. Its $V_{max}$ values for phenylalanine and tyrosine were 1.77 and $0.47{\mu}mol$/ min/mg protein, respectively, and its $K_m$ values were $4.77{\times}10^{-4}$ and $4.37{\times}10^{-4}\;M$, respectively. PAL showed an optimal pH at 8.5, corresponding to the average pH range of the small intestine. It showed no loss of activity at $-80^{\circ}C$ for 5 months and possessed 93.4% of its activity under $4^{\circ}C$ for 4 wks. PAL was susceptible to chymotrypsin digestion and, to a lesser extent, to trypsin, elastase, carboxypeptidase A, and B. The trypsin and chymotrypsin cleaving sites were mutated to investigate protection from pancreatic digestion and the specific activities of these mutants were evaluated. The six mutants displayed low specific activities compared to the wild-type, suggesting that the primary trypsin and chymotrypsin cleaving sites may be essential for catalytic reaction. The PAL mutants could therefore be applied as a pretreatment modality without susceptibility to proteolytic attack, however, additional modification for enhancing enzymatic activity is needed to reduce the Phe levels effectively.

Change of the Result Value by the Amount of Samples in CA 19-9 (CA 19-9 검체량에 따른 농도의 변화)

  • Chang, Hyun-Yeong;Park, Hee-Won;You, Tae-Min;Noh, Kyung-Woon;Kim, Hyun-Joo
    • The Korean Journal of Nuclear Medicine Technology
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    • v.12 no.1
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    • pp.66-73
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    • 2008
  • Purpose: CA 19-9 need to examine a number of sample volume, and the postwar 200 U/ml concentration hook effect appears slight. Thus, the antibody-antigen reaction, and by reducing the amount of (sample volume), they can hook effect to minimize the impact of the sample volume and relevance know, I saw the hook effect. In addition, the current maximum of using the standard concentration of the reagent in 240 U/ml increase more than the standard concentration can be seen knows. Material and Method: 5 U/ml and under, make a few low concentration of serum pool from the high concentration of the sample hook effect together with a standard concentration of about 500 to meet the production. The reagents used in experiments are currently using SNUH NM experiment. Orignal method along with the experiment is to 25 ul sample volume (1 / 4), 50 ul (1 / 2), 100 ul (Orignal method) in the experiment. My greatest concentration of the reagent concentration of approximately two times the standard concentration of production. When was the last to make the first experiment, as measured by the standard concentration after that. The new inspection information through a standard solution modified by entering values in them. Results: 100 ul, and to apply the new standard concentration y = 1.3021x - 10.97, $R^2$ = 0.9844. Overall, the results showed a similar orignal method. Because of the concentration in the value of more than 240 U/ml, but it is an overall value that can be made out of a similar value When I put the 50 ul y = 1.045x + 9.5861, $R^2$ = 0.9428. Overall orignal method and the results of a similar value. 50 ul, and to apply the new standard concentration y=1.2006x+11.252, $R^2$=0.9423. Showing a slightly lower value compared with orignal method. Because of the concentration in the value of more than 240 U/ml, but it is an overall value that can be made out of a similar value. When I put the man 25 ul y=0.6012x+24.755, $R^2$=0.4033. Results showed that very small amounts of sample are insecure inside and showed a lower middle cpm orignal method and showed a lot of mismatched. Conclusions: 25 ul of the sample volume is not possible to use the instability had, when I put the 50 ul of the orignal method can be used to show a similar concentration. The new values are slightly lower concentration, The new values are slightly lower concentration, concentration, which are likely due to the lack of data has had a little gap between the sample showed 80 to 200 U/ml additional experiments seem to do. Apply a new 100 ul concentration values are applied to a large crowd is not even in sight. But this way the concentration of 100 to more 400 U/ml gather further experiments should possible adds.

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Agronomic Traits of Soybean Breeding Lines with Low Stachyose and Raffinose Contents (Stachyose 및 Raffinose 저함량 콩 선발계통의 농업적 형질)

  • Ha, Do Su;Moon, Jin Young;Choi, Sang Woo;Shim, Sang In;Kim, Min Chul;Chung, Jong Il
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.62 no.2
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    • pp.143-148
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    • 2017
  • Soybean [Glycine max (L.) Merr.] seed is an important dietary source of protein, oil, carbohydrates, isoflavones, and other nutrients for humans and animals. Raffinose and stachyose are the main antinutritional factors in soybean seed. They are carbohydrates belonging to the raffinose family of oligosaccharides, which are not readily digested in humans and cause flatulence or diarrhea. The genetic reduction of the raffinose and stachyose contents in mature soybean seeds will improve the nutritional value of soybean. The objective of this research was to evaluate agronomic traits with 10 $F_6$ strains selected from breeding populations derived from a cross among seven parents. The contents of raffinose and stachyose in mature seeds were detected by high-performance liquid chromatography. Agronomic traits such as flower color, flowering date, harvesting date, lodging, plant height, seed coat color, hilum color, 100 seed weight, and yield were evaluated. Ten intermediate parents showed low raffinose and stachyose contents. The intermediate parent 883-1 had a small seed size, six intermediate parents (15A1, 15D1, RS-5, RS-33, RS-64, and RS-70) had a medium seed size, and two intermediate parents (14G20 and RS-21) had a large seed size. The intermediate parent RS-21 had a black seed coat and a green cotyledon. Four intermediate parents (883-1, 14G20, RS-5, and RS-21) had elite agronomic traits. The new intermediate parents developed through this study will be used to develop improved soybean cultivars with low contents of raffinose and stachyose.

Seasonal Changes of Copper and Manganese Content in the Raw Bovine Milk in Korea (국내산 원유 중의 구리 및 망간 함량의 계절적 변화)

  • Kwak Byung-Man;Jun Yeun-Myoung;Kim Kang-Seob;Lee Ki-Woong;Ahn Jang-Hyuk;Chang Chi-Hoon
    • Food Science of Animal Resources
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    • v.26 no.2
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    • pp.241-244
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    • 2006
  • This study was conducted by using the ICP-AES (Inductively Coupled Plasma - Atomic Emission Spectrometry) to investigate the seasonal changes of copper and manganese content in the raw bovine milk. From July 2003 to June 2004, the milk samples were collected from the two geographical locations, Chungcheong-do and Jeolla-do, in Korean peninsula. Copper (Cu) content was determined in the range of min. 10.10 to max. 21.00 ${\mu}g$/100 mL (13.79 ${\mu}g$/100 mL) in spring, 5.06 to 15.41 (10.17) in summer, 5.04 to 19.70 (10.70) in autumn and 6.96 to 17.90 (12.11) in winter. For manganese (Mn), 3.00 to 8.30 (4.87) in spring, 2.30 to 6.44 (3.75) in summer, 2.81 to 6.04 (3.82) in autumn and 2.25 to 9.02 (4.48) in winter. Those data have shown that levels of copper and manganese was relatively constant but higher in Spring than other seasons, suggesting that the levels were not affected by seasons and different locations in Korea.

The Study on the Methylmercury Analysis and the Monitoring of Total Mercury and Methylmercury in Fish (어류 중 메틸수은 분석법 확립 및 모니터링)

  • Kim, Hee-Yun;Chung, So-Young;Sho, You-Sub;Oh, Geum-Soon;Park, Seong-Soo;Suh, Jung-Hyuk;Lee, Eun-Ju;Lee, Yoon-Dong;Choi, Woo-Jeong;Eom, Ji-Yoon;Song, Min-Soo;Lee, Jong-Ok;Woo, Gun-Jo
    • Korean Journal of Food Science and Technology
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    • v.37 no.6
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    • pp.882-888
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    • 2005
  • Procedure for analysis of methylmercury in fish was developed, involving addition of HCl, extraction with toluene, and clean-up using L-cystein solution. Obtained extract is analyzed by gas chromatography with electron capture detector using Ulbon HR-Thermon-Hg column. Detection limit and recovery of the method were 0.005mg/kg (expressed as Hg), 98-107 (103%), respectively. Total mercury and methylmercury concentrations in 175 commercial fish samples ranged from [mean-max (mean), unit: mg/kg]: 0.014-1.200 (0.270) and 0.006-0.901 (0.168) in tuna-fish, 0.020-0.934 (0.323) and 0.012-0.553 (0.149) in martin-fish, 0.082-0.782 (0.391) and 0.040-0.436(0.201) in shark, 0,023-0.031 (0.026) and 0,013-0.018 (0.015) in salmon, 0.098-0.193 (0.133) and 0.031-0.015(0.090) in tilefish, and 0,031-0.214 (0.089) and 0.016-0.093 (0.042) in canned tuna respectively. No sample of analyzed fish exceeded 1.0mg/kg wet wt., limit for methylmercury established by Codex. In all species examined, estimated weekly intake was lower than Provisional Tolerable Weekly Intake recommended by the JECFA (the Joint FAO/WHO Expert Committee on Food Additives).

Evaluating applicability of metal artifact reduction algorithm for head & neck radiation treatment planning CT (Metal artifact reduction algorithm의 두경부 CT에 대한 적용 가능성 평가)

  • Son, Sang Jun;Park, Jang Pil;Kim, Min Jeong;Yoo, Suk Hyun
    • The Journal of Korean Society for Radiation Therapy
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    • v.26 no.1
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    • pp.107-114
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    • 2014
  • Purpose : The purpose of this study is evaluation for the applicability of O-MAR(Metal artifact Reduction for Orthopedic Implants)(ver. 3.6.0, Philips, Netherlands) in head & neck radiation treatment planning CT with metal artifact created by dental implant. Materials and Methods : All of the in this study's CT images were scanned by Brilliance Big Bore CT(Philips, Netherlands) at 120kVp, 2mm sliced and Metal artifact reduced by O-MAR. To compare the original and reconstructed CT images worked on RTPS(Eclipse ver 10.0.42, Varian, USA). In order to test the basic performance of the O-MAR, The phantom was made to create metal artifact by dental implant and other phantoms used for without artifact images. To measure a difference of HU in with artifact images and without artifact images, homogeneous phantom and inhomogeneous phantoms were used with cerrobend rods. Each of images were compared a difference of HU in ROIs. And also, 1 case of patient's original CT image applied O-MAR and density corrected CT were evaluated for dose distributions with SNC Patient(Sun Nuclear Co., USA). Results : In cases of head&neck phantom, the difference of dose distibution is appeared 99.8% gamma passing rate(criteria 2 mm / 2%) between original and CT images applied O-MAR. And 98.5% appeared in patient case, among original CT, O-MAR and density corrected CT. The difference of total dose distribution is less than 2% that appeared both phantom and patient case study. Though the dose deviations are little, there are still matters to discuss that the dose deviations are concentrated so locally. In this study, The quality of all images applied O-MAR was improved. Unexpectedly, Increase of max. HU was founded in air cavity of the O-MAR images compare to cavity of the original images and wrong corrections were appeared, too. Conclusion : The result of study assuming restrained case of O-MAR adapted to near skin and low density area, it appeared image distortion and artifact correction simultaneously. In O-MAR CT, air cavity area even turned tissue HU by wrong correction was founded, too. Consequentially, It seems O-MAR algorithm is not perfect to distinguish air cavity and photon starvation artifact. Nevertheless, the differences of HU and dose distribution are not a huge that is not suitable for clinical use. And there are more advantages in clinic for improved quality of CT images and DRRs, precision of contouring OARs or tumors and correcting artifact area. So original and O-MAR CT must be used together in clinic for more accurate treatment plan.

An Experimental Study on Flow Characteristics for Optimal Spacing Suggestion of 45° Upward Groynes (45° 상향수제의 적정 간격 제시를 위한 흐름특성 실험 연구)

  • Kim, Sung Joong;Kang, Joon Gu;Yeo, Hong Koo
    • Journal of Korea Water Resources Association
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    • v.47 no.5
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    • pp.459-468
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    • 2014
  • Groyne to control the direction and velocity of flow in rivers is generally installed for the purpose of protecting riverbanks or embankments from erosion caused by running water. In particular, as interest in river restoration and natural river improvement increases, groynes are proposed as a key hydraulic structure for local flow control and riparian habitat establishment. Groynes are installed mainly in groups rather than as individual structures. In case of groynes installed as a group, flow around the groynes change according to spacing in between the groynes. Therefore, groyne spacing is regarded as the most important factor in groyne design. This study aimed at examining changes of flows around and within the area of groynes that take place according to the spacing of groynes installed in order to propose the optimal spacing for upward groynes. To examine flow characteristics around groynes, this study looked at flows in main flow area and recirculation flow area separately. In main flow area, it examined the impact of flow velocity increasing as a result of conveyance reduction that is exerted on river bed stability in relation to changes in the maximum flow velocity according to installation spacing. As a factor causing impacts on scouring and sedimentation within the area of groynes, recirculation flow in the groyne area can lead problems concerning flow within the area and stability of embankment. As for recirculation area, an analysis was conducted on the scale of rotational flow and the flow around embankment that exerts impacts on stability of the embankment. In addition, a comparative analysis was carried with reference to changes of the central point of rotational flow that occur within the area of groynes. As a result of compositely examining the results, the appropriate installation spacing is proposed as min. four times-max. six times considering a decrease in flow velocity according to the installation of upward groynes, river bed stability and stability of embankments against counterflow within the area of groynes.

The Difference of Interpretations of Cardiopulmonary Exercise Testing According to Interpretative Algorithms and Exercise Methods (분석 알고리즘과 운동방법에 따른 Exercise Test 결과의 차이)

  • Park, Jae-Min;Kim, Sung-Kyu
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.1
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    • pp.42-51
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    • 2001
  • Background : Recently, cardiopulmonary exercise testing (CPX) has become a popular diagnostic method for differentiating the main cause of exertional dyspnea or exercise limitation. We evaluated the difference in the CPX results according to interpretative algorithms and the methals of exercise in Korea. Method : Sixty-six patients with chronic lung disease and 48 adults with dyspneic symptoms, but with no abnormalities in a spirometry performed symptom limited CPX, were included in this study. The results were interpreted using both Wasserman's(WA) and Eschenbacher's algorithm (EA), and a comparison between both algorithms was made. Thirty-three healthy medical students performed the CPX with a cycle ergometer and treadmill. The results were interpreted with EA and the concurrence in interpretations was evaluated accord ing to the methods of exercise. Results : 1. In patients with chronic lung disease, the overall concordance rate between the two algorithms was 63.6%. The concordance rates waw 69.8% in patients with obstructive, 25.0% in those with restrictive, and 66.7% in those with mixed pulmonary insufficiency. In patients with dyspneic symptoms but normal findings in resting spirometry, the concordance rate was 60.4%. 2. In healthy medical students, in results inter preted with EA, the concordance rate between the cycle ergometer and treadmill exercise was 25.0%. Conclusion : Both interpretative algorithms and methods of exercise may affect the CPX results. In using CPX as a diagnostic test for the causes of dyspnea in the Korean population. the interpretative algorithms and method of exercise need to be standardized, and a predictive $VO_2$max equation needs to be established.

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Comparison of Anticancer and Immuno-Modulatory Activities in the different parts of the Acer mono Max. and Acer okamotoanum (고로쇠와 우산고로쇠 나무의 부위별 항암 및 면역조절능 비교)

  • Qadir, Syed Abdul;Kim, Cheol-Hee;Kwon, Min-Chul;Lee, Hak-Ju;Kang, Ha-Young;Choi, Don-Ha;Lee, Hyeon-Yong
    • Korean Journal of Medicinal Crop Science
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    • v.15 no.6
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    • pp.405-410
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    • 2007
  • This study was performed to investigate anticancer activities and immuno modulatory activities in the several parts of the A. mono and A. okamotoanum. The cytotoxicity of 1 $mg/m{\ell}$ of the water extracts on normal human lung cell(HEL299) was < 19.5%. The anticancer activity of all extracts were increased in over 55% against AGS (stomach adenocarcinoma), A549 (lung adenocarcinoma), Hep3B (liver adenocarcinoma, and MCF-7 (breast adenocarcinoma) cells. The growth of human immune B and T cells was improved of A. mono and A. okamotoanum in adding 1.0 $mg/m{\ell}$ concentration. The secretion of the IL-6 and $TNF-{\alpha}$ of human immune B and T cells was increased with all extracts of A. mono and A. okamotoanum. All extracts of. A. mono and A. okamotoanum increased NK cell growth. The results showed that the barks and woods extracts of A. mono and A. okamotoanum had useful biological activities. In addition, bark of A. okamotoanuim showed the highest anticancer and immune activities.

Inheritance of Kunitz Trypsin Inhibitor and P34 Protein in Soybean Seed (콩 종자에서 쿠니츠트립신인히비터와 P34 단백질의 유전)

  • Han, Eun-Hui;Sung, Mi-Kyung;Baek, Woon-Jang;Shim, Sang-In;Kim, Min-Chul;Chung, Jong-Il
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.57 no.1
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    • pp.78-82
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    • 2012
  • Soybean [$Glycine$ $max$ (L.) Merr.] protein is a high quality source for food and feed. But, antinutritional factors in the raw mature soybean are exist. Kunitz trypsin inhibitor (KTI) protein is a main antinutritional factor in soybean seed. Also, P34 protein, referred as $Gly$ $m$ Bd 30K, has been identified as a predominant immunodominant allergen. Genetic relationship between KTI protein and P34 protein could be useful in soybean breeding program for the genetic elimination or reduction of these factors. The objective of this study was to determine the independent inheritance or linkage between KTI protein and P34 protein in soybean seed. A total of 479 $F_2$ seeds were obtained from the cross of 07B1 and PI567476 parents. KTI protein and relative amount of P34 protein were analysed from $F_2$ seeds harvested from the F1 plants by using SDS-PAGE and Western blot analysis. The segregation ratios of 3 : 1 for KTI protein (353 KTI protein present : 126 KTI protein absent) and relative amount of P34 protein (363 normal amount of P34 protein : 116 low amount of P34 protein). The segregation ratio of 3 : 1 suggested that KTI protein and relative amount of P34 protein in mature soybean seed were controlled by a single major gene. The segregation ratios of 9 : 3 : 3 : 1 (266 KTI protein present, normal amount of P34 protein: 88 KTI protein present, low amount of P34 protein: 102 KTI protein absent, normal amount of P34 protein: 23 KTI protein absent, low amount of P34 protein) and Chi-square value (${\chi}^2$=3.31, P=0.346) were observed in $F_2$ seeds. This data showed that KTI protein was inherited independently with relative amount of P34 protein in soybean. These results will be helpful in breeding program for selecting the line with lacking KTI protein and reduced amount of P34 protein in soybean.