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Protein Kinase Modulates the $GABA_c$ Currents in Cone-horizontal Cell Axon-terminals Isolated from Catfish Retina

  • Paik, Sun-Sook;Lee, Sung-Jong;Jung, Chang-Sub;Bai, Sun-Ho
    • Proceedings of the Korean Biophysical Society Conference
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    • 1999.06a
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    • pp.54-54
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    • 1999
  • Protein kinase modulation of gamma-aminobutyric acid C (GABA$_{c}$) currents in freshly dissociated catfish retinal cone-horizontal cell axon-terminals was studied under voltage clamp with the use of the whole cell patch-clamp technique. Responses to pulses of GABA were monitored in intracellular application of adenosin 3',5'-cycle monophophate (cAMP)-dependent protein kinase (PKA) and protein kinase C (PKC) activators, and their inhibitors or inactive analogues.(omitted)d)

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CUEDC2, CUE Domain Containing Protein 2, Associates with Kinesin-1 by Binding to the C-Terminus of KIF5A (CUE 도메인 포함 단백질인 CUEDC2는 KIF5A의 C-말단과 결합을 통하여 Kinesin-1와 결합)

  • Myoung Hun Kim;Se Young Pyo;Young Joo Jeong;Sung Woo Park;Mi Kyoung Seo;Won Hee Lee;Sang-Hwa Urm;Mooseong Kim;Jung Goo Lee;Dae-Hyun Seog
    • Journal of Life Science
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    • v.33 no.11
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    • pp.868-875
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    • 2023
  • Kinesin-1 is a motor protein identified as the first member of the kinesin superfamily (KIF), which plays a role in intracellular cargo transport by acting as microtubule-dependent motor proteins within cells. Kinesin-1 consists of two heavy chains (KHCs, also known as KIF5s) and two light chains (KLCs). The 93 amino acids in the carboxyl (C)-terminal tail region of KIF5A are not homologous to the C-terminal tail region of KIF5B or the C-terminal tail region of KIF5C. In this study, we used a yeast two-hybrid screen to identify the binding proteins that interacted with the C-terminal region of KIF5A. We found an association between KIF5A and CUE domain containing 2 (CUEDC2), which is proposed to function as an adaptor protein involved in ubiquitination pathways and protein trafficking. CUEDC2 bound to the C-terminal region of KIF5A and did not interact with KIF5B (the motor of kinesin-1), KIF3A (the motor of kinesin-2), or kinesin light chain 1 (KLC1). KIF5A specifically bound to the C-terminal region of CUEDC2. Furthermore, KIF5A did not interact with another isoform: CUEDC1. In addition, glutathione S-transferase (GST) pull-downs showed that KIF5A directly bound GST-CUEDC2 but did not interact with GST-CUEDC1 and GST alone. When myc-KIF5A and EGFP-CUEDC2 were co-expressed in HEK-293T cells, CUEDC2 co-immunoprecipitated with kinesin-1, and myc-KIF5A and FLAG-CUEDC2 colocalized in the cells. These results suggest that in intracellular cargo transport by kinesin-1, CUEDC2 serves as an adaptor protein connecting kinesin-1 and cargo by binding to KIF5A.

The Fundamental Studies of Heat Treatment Class in Domestic Infant Formula (국산 조제분유의 열처리 등급에 관한 기초 연구)

  • 박영희
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.27 no.3
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    • pp.495-500
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    • 1998
  • To compare the extent of heat treatment in domestic infant formula, pH,titratable acidity, undenatured whey protein contents, HMF contents and protein-reducing substances of three commercial products (A, B, C) were measured. The pH of B products was lowest and the titratable acidity of B product was highest. The contents of undenatured whey protein per 100ml serum were 0∼30mg(A products), 90∼130mg(B products)and 80∼90mg(C products), respectively. Distinct differences of undenatured whey protein contents according to the manufacturer and infat's stage in age could be observed. The HMF contents of tested products showed 10.9∼21.5umol/L and B-2 product(B products for the second stage of 5∼9 month) was the highest among tested products. The protein-reducing substances showed 4.46∼9.50mg K4Fe(CN)6/100ml serum nd B-2 product was the highest among tested products.

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Performance of Broiler Chickens Fed Low Protein, Limiting Amino Acid Supplemented Diets Formulated Either on Total or Standardized Ileal Digestible Amino Acid Basis

  • Kumar, C. Basavanta;Gloridoss, R.G.;Singh, K.C.;Prabhu, T.M.;Suresh, B.N.
    • Asian-Australasian Journal of Animal Sciences
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    • v.29 no.11
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    • pp.1616-1624
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    • 2016
  • The aim of present experiment was to investigate the effect of protein reduction in commercial broiler chicken rations with incorporation of de-oiled rice bran (DORB) and supplementation of limiting amino acids (valine, isoleucine, and/or tryptophan) with ration formulation either on total amino acid (TAA) or standardized ileal digestible amino acids (SIDAA). The experimental design consisted of $T_1$, TAA control; $T_2$ and $T_3$, 0.75% and 1.5% protein reduction by 3% and 6% DORB incorporation, respectively by replacing soybean meal with supplemental limiting amino acids to meet TAA requirement; $T_4$, SIDAA control, $T_5$ and $T_6$, 0.75% and 1.5% protein reduction by DORB incorporation (3% and 6%) with supplemental limiting amino acids on SIDAA basis. A total of 360 dold fast growing broiler chicks (Vencobb-400) were divided into 36 homogenous groups of ten chicks each, and six dietary treatments described were allocated randomly with six replications. During 42 days trial, the feed intake was significantly (p<0.05) reduced by TAA factor compared to SIDAA factor and protein factor significantly (p<0.05) reduced the feed intake at 1.5% reduction compared to normal protein group. This was observed only during pre-starter phase but not thereafter. The cumulative body weight gain (BWG) was significantly (p<0.05) reduced in TAA formulations with protein step-down of 1.5% ($T_3$, 1,993 g) compared to control ($T_1$, 2,067 g), while under SIDAA formulations, BWG was not affected with protein reduction of 1.5% ($T_6$, 2,076 g) compared to $T_4$ (2,129 g). The feed conversion ratio (FCR) was significantly (p<0.05) reduced in both TAA and SIDAA formulations with 1.5% protein step-down ($T_3$, 1.741; $T_6$, 1.704) compared to respective controls ($T_1$, 1.696; $T_4$, 1.663). The SIDAA formulation revealed significantly (p<0.05) higher BWG (2,095 g) and better FCR (1.684) compared to TAA formulation (2,028 g; 1.721). Intake of crude protein and all limiting amino acids (SID basis) was higher in SIDAA group than TAA group with resultant higher nitrogen retention (4.438 vs 4.027 g/bird/d). The nitrogen excretion was minimized with 1.5% protein reduction (1.608 g/bird) compared to normal protein group (1.794 g/bird). The serum uric acid concentration was significantly reduced in $T_3$ (9.45 mg/dL) as compared to $T_4$ (10.75 mg/dL). All carcass parameters were significantly (p<0.05) higher in SIDAA formulation over TAA formulation and 1.5% protein reduction significantly reduced carcass, breast and thigh yields. In conclusion, the dietary protein can be reduced by 0.75% with TAA formulation and 1.5% with SIDAA formulation through DORB incorporation and supplementation of limiting amino acids and among formulations, SIDAA formulation was better than TAA formulation.

Effects of a High Protein Diet and Aerobic Exercise on Body Weight Changes and Blood Lipids in Slightly Overweight Women

  • Suh, Kyoung Yeo;Lee, Dae Taek
    • Nutritional Sciences
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    • v.8 no.1
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    • pp.28-34
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    • 2005
  • To examine the combined effects of a high-protein diet and aerobic exercise on body weight and composition and blood lipid profiles in overweight women, 30 young women were recruited and placed into three groups: The high-protein diet and exercise group (HPE), the exercise-only group (EXO) and the control group (CON) (30$\pm$3%, 27$\pm$2%, and 29$\pm$3% body fat, respectively) for an 8-week experimental period. Daily diet included 25% isolated soybean protein (>90% protein, approximately 400 kcal) combined with each subject s usual diet for the HPE group. The exercise program consisted of aerobic-type exercises undertaken >3 times/wk and for>30 min/session at 50-60% of maximal capacity. Physical fitness, body composition, serum total cholesterol (TC), high-density lipoprotein-cholesterol (HDL-C), triglycerides (TG) and glucose were measured before and after the experiment. Maximal aerobic capacity increased by the end of experiment in both the HPE (from 27.2$\pm$3.5 to 35.l$\pm$5.9 ml/kg/min, p<0.01) and EXO (from 30.3$\pm$5.4 to 33.8$\pm$3.8 mㅣ/kg/min, p<0.05) groups. Percent body fat decreased by 3.3% (p<0.01) in the HPE group and by 1.5% (p<0.05) in the EXO group by the end of the experiment, but not in the CON group. Lower back strength and agility increased only in the HPE group. In the HPE group, TC decreased from 168$\pm$20 to 155$\pm$18 mg/dL and HDL-C increased from 57$\pm$l0 to 61$\pm$9 mg/dL in HPE (p<0.01). But TC and HDL-C did not change in the EXO and CON groups. TG and glucose did not vary among the groups. Although the EXO group showed a similar outcome to that of the HPE group, a favorable change in body composition and blood lipids as well as an improvement in aerobic capacity was more marginal in the latter group.

Identification of 3'-Hydroxymelanetin and Liquiritigenin as Akt Protein Kinase Inhibitors

  • Yang Hye-Young;Lee Hong-Sub;Ko Jong-Hee;Yeon Seung-Woo;Kim Tae-Yong;Hwang Bang-Yeon;Kang Sang-Sun;Chun Jae-Sun;Hong Soon-Kwang
    • Journal of Microbiology and Biotechnology
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    • v.16 no.9
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    • pp.1384-1391
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    • 2006
  • The signal transduction system is one of the most important devices involved in maintaining life, and many protein kinases are included in the cellular signal transduction system. Finding a protein kinase inhibitor is very valuable, as it can be used to study cell biology and applied to pharmaceuticals. For the efficient and rapid screening of protein kinase inhibitors, two assay systems were combined; the nonradioactive protein kinase assay system that uses an FITC-labeled IRS-2 peptide and the cell-based paper disc assay system that uses Streptomyces griseus as the indicator strain. Among 330 kinds of herb extracts tested, the extract of Dalbergia odorifera exhibited the strongest inhibitory activity in the two assay systems and was selected for further isolation. Based on solvent extraction and many steps of chromatography, seven compounds were finally separated to homogeneity and their structures determined by $^{1}H$ and $^{13}C$ NMR spectroscopies. Four were to be flavonoids and identified as butin ($C_{15}H_{12}O_5$, Mw=272.07), 3'-hydroxymelanetin ($C_{16}H_{12}O_6$, Mw=300.06), liquiritigenin ($C_{15}H_{12}O_4$, Mw=256.07), and 2'-hydroxyformononetin ($C_{16}H_{12}O_{5}$, Mw=284.07). 3'-Hydroxymelanetin inhibited the phosphorylation of the GSK3 protein by Akt to 37% at a concentration of $10{\mu}g/ml$ and showed the strongest cytotoxicity ($ED_{50}<50{\mu}g/ml$) against the human cancer cell line HCT116. Under the same conditions, liquiritigenin also inhibited the phosphorylation of GSK3 by Akt to 26%, and its cytotoxicity against the HCT116 cell line was lower than $100{\mu}g/ml$.

Overexpression and Purification of PreS Region of Hepatitis B Virus Antigenic Surface Protein adr Subtype in Escherichia coli

  • Abbas, Naaz;Ahmad, Aftab;Shakoori, Abdul Rauf
    • BMB Reports
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    • v.40 no.6
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    • pp.1002-1008
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    • 2007
  • PreS domain of Hepatitis B virus (HBV) surface antigen is a good candidate for an effective vaccine as it activates both B and T cells besides binding to hepatocytes. This report deals with overexpression and purification of adr subtype of surface antigen that is more prevalent in Pakistan. PreS region, comprising 119 aa preS1 region plus a 55 aa preS2 region plus 11 aa from the N-terminal S region, was inserted in pET21a+ vector, cloned in E. coli $DH5\alpha$ cells and expressed in E. coli BL21 codon+ cells. The conditions for over expression were optimized using different concentrations of IPTG (0.01-5 mM), and incubating the cells at different temperatures (23-$41^{\circ}C$) for different durations (0-6 h). The cells were grown under the given optimized conditions (0.5 mM IPTG concentration at $37^{\circ}C$ for 4 h), lysed by sonication and the protein was purified by ion exchange chromatography. On the average, 24.5 mg of recombinant protein was purified per liter of culture. The purified protein was later lyophilized and stored at $-80^{\circ}C$.

Application of Temperature Gradient Gel Electrophoresis To cAMP Receptor Protein (온도 기울기 전기영동장치의 CAMP 수용성 단백질에 응용)

  • Gang, Jong-Back;Cho, Hyun-Young
    • Journal of Life Science
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    • v.14 no.2
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    • pp.309-314
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    • 2004
  • Cyclic AMP receptor protein (CRP) is involved in the transcriptional regulation of more than 100 genes in E. coli. CRP dimer is converted into active form via the sequential conformation change of cAMP binding pocket, hinge region and HTH DNA binding motif by binding of cAMP. The temperature gradient gel electrophoresis (TGGE) was applied to CRP protein to know whether it was an efficient technique to study the conformational transitions and the thermal stability. TGGE showed the unfolding process of wild-type and S83G CRP proteins with the temperature gradient set from 29 to 71$^{\circ}C$ on nondenaturing polyacrylamide gel. Melting temperature (Tm) was 57$\pm$1 and 55$\pm$1$^{\circ}C$ for wild-type and S83G CRP, respectively in acidic buffer[89.8 mM Glycine and 24 mM Boric acid (pH 5.8)].

Molecular Cloning and Expression of a Novel Cuticle Protein Gene from the Chinese Oak Silkmoth, Antheraea pernyi

  • Kim Bo Yeon;Park Nam Sook;Jin Byung Rae;Kang Pil Don;Lee Bong Hee;Seong Su Il;Hwang Jae Sam;Chang Jong Su;Lee Sang Mong
    • International Journal of Industrial Entomology and Biomaterials
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    • v.10 no.1
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    • pp.11-17
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    • 2005
  • In our research to identify gene involved in the cuticle protein, we cloned a novel cuticle protein gene, ApCP15.5, from the Chinese oak silkmoth, Antheraea pernyi, larvae cDNA library. The gene encodes a 149 amino acid polypeptide with a predicted molecular mass of 15.5 kDa and a pI of 9.54. The ApCP15.5 contained a type-specific consensus sequence identifiable in other insect cuticle proteins and the deduced amino acid sequence of the ApCP15.5 cDNA is most homologous to Tenebrio molitor-C1B ($43\%$ protein sequence identity), followed by Locusta migratoria-76 ($42\%$ protein sequence identity). Northern blot and Western blot analyses revealed that the ApCP15.5 showed the epidermis-specific expression. The expression profile of ApCP15.5 indicated that the ApCP15.5 mRNA expression was detected in the early stages after larval ecdysis and larval-pupal metamorphosis, and its expression level was most significant on the first day of larval ecdysis and pupal stage. The ApCP15.5 was expressed as a 15.5 kDa polypeptide in baculovirus-infected insect cells.

Effect of Partial Replacement of Fish Meal by New Squid Sepia esculenta Liver Powders on the Growth and Body Composition of Juvenile Olive Flounder Paralichthys olivaceus (넙치(Paralichthys olivaceus) 치어 사료에 오징어(Sepia esculenta) 간분말 신제품에 대한 어분대체 효과)

  • MoonLee, Hae-Young;Choi, Se-Min;Ji, Ho-Seok
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.45 no.2
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    • pp.132-138
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    • 2012
  • An 8-week feeding experiment was conducted to evaluate three types of squid Sepia esculenta liver powder (SLP) as a dietary protein source replacing fish meal (FM) in juvenile olive flounder Paralichthys olivaceus. To replace FM, six experimental diets were formulated with three types (A, B, C) of SLP at the 5 and 10%: SLP-A5, SLP-A10, SLP-B5, SLP-B10, SLP-C5, and SLP-C10. One control diet contained 100% FM as the main protein source and another was a commercial diet (Com). Fish with an average body weight of $22.8{\pm}0.4$ g ($mean{\pm}SD$) were allocated randomly to aquaria in groups of 20 fish and fed the experimental diets in triplicate to satiation. The weight gain (WG) and specific growth rate (SGR) of fish fed the SLP-C10 diet were lower than those of fish fed the FM and SLP-B5 diets. No significant difference was observed in the WG and SGR among fish fed the diets other than SLP-C10. The feed efficiency (FE) and protein efficiency ratio (PER) of fish fed each SLP diet did not differ from those fed the control diet. However, fish fed SLP-C5 and SLP-C10 had a lower FE and PER than the fish fed commercial, SLP-A5 and SLP-B5 diets. Each SLP diet except for SLP-C10 could replace up to 10% of FM for juvenile olive flounder. The results of this experiment provide information that will assist in formulating an inexpensive practical diet containing SLP for juvenile olive flounder.