• Title/Summary/Keyword: Leu-7

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Purification and Characterization of Substance P-related Peptide from the Body of the African Lungfish, Protopterus dolloi

  • Kim, Chan-Hee;Kim, Eun-Jung;Go, Hye-Jin;Lee, Hyung-Ho;Hong, Yong-Ki;Kim, Hyung-Rak;Chung, Joon-Ki;Park, Jang-Su;Muneoka, Yojiro;Park, Nam-Gyu
    • Bulletin of the Korean Chemical Society
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    • v.27 no.7
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    • pp.1015-1019
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    • 2006
  • The peptide with structural similarity to mammalian substance P (M-SP) has been isolated from extract of the body of the African lungfish, Protopterus dolloi, using the rectum of the newt as the bioassay system. The primary structure of the SP-related peptide was identified as Lys-Pro-Arg-Pro-Asp-Gln-Phe-Tyr-Gly-Leu-Met-NH2 (L-SP) and contained four substitutions ($Lys^{1}\rightarrow $ Arg, $Arg^{3}\rightarrow$ Lys, $Asp^{5}\rightarrow$ Gln, and $Tyr^{8}\rightarrow$ Phe) compared with M-SP; this structure is identical to that of the peptide isolated from the gut of the Australian lungfish. Circular dichroism spectra showed that L-SP had an unordered structure in the buffer solution and phospholipid bilayers. This peptide was found to have an excitatory effect on rectal muscle tissues of newt, quail, and fish. L-SP also had a more potent vasodilatory effect on the guinea-pig aorta than that of M-SP. The identification of the peptide provides evidence that SP family, hitherto confined to mammals, have a widespread occurrence in lungfish.

Purification and Characterization of a Thermostable Xylose (Glucose) Isomerase from Streptomyces chibaensis J-59

  • Joo, Gil-Jae;Shin, Jae-Ho;Heo, Gun-Young;Kwak, Yun-Young;Choi, Jun-Ho;Rhee, In-Koo
    • Journal of Applied Biological Chemistry
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    • v.44 no.3
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    • pp.113-118
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    • 2001
  • Xylose (glucose) isomerase was purified to homogeneity from cell-extracts of Streptomyces chibaensis J-59 via ammonium sulfate precipitation followed by chromatography on DEAE-cellulose, and gel filtration on Sephacryl S-300. The purified enzyme is a homotetramer with a native molecular mass of 180 kDa and a subunit molecular mass of 44 kDa. The amino acid N-terminal sequence of glucose isomerase from S. chibaensis J-59 was determined to be Ser-Tyr-Gln-Pro-Thr-Pro-Glu-Asp-Arg-Phe-Thr-Phe-Gly-Leu. The first 14 amino acids of the N-terminal sequence of the enzyme showed strong analogies with N-terminal sequences of glucose isomerase produced by other Streptomyces spp. The optimum pH and temperature for activity were 7.5 and 85, respectively. The purified enzyme required $Mg^{2+}$, $Co^{2+}$, and $Mn^{2+}$ for the activity, $Mg^{2+}$ being the most effective. The enzyme was not inhibited by $Ca^{2+}$, but was inhibited by $Hg^{2+}$, $Ag^+$, and $Cu^{2+}$. The $K_m$, $V_{max}$, and $k_{cat}$ values of S. chibaensis J-59 isomerase for glucose were 83 mM, 40.9 U/mg, and $1,843min^{-1}$, respectively. In the presence of $Co^{2+}$, cell-free enzymes retained 100% without loss of activities by the heat-treatment at $70^{\circ}C$ for 7 days. The enzyme retained 50% residual activity after heating at $85^{\circ}C$ for 13.5 h, at $90^{\circ}C$ for 126 min. The enzyme is more thermostable than any other glucose isomerases of Streptomyces spp.

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Inhibition of Aminopeptidase N by Two Synthetic Tripeptides

  • Chung, Myung Chul;Hyo Kon Chun;Ho Jae Lee;Choong Hwan Lee;Su Il Kim;Yung Hee Kho
    • Journal of Microbiology and Biotechnology
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    • v.6 no.1
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    • pp.7-11
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    • 1996
  • MR-387Al (ARPA-Val-Pro) and A2 (AHPA-Val-Hyp) were prepared as aminopeptidase N inhibitors through the synthesis of peptide MR-387A and B analogues which contained 3-amino-2-hydroxy-4-phenyl butanoic acid (ARPA) as a zinc-chelating moiety. They are competitive inhibitors of aminopeptidase N with inhibition constants(Ki) of 4.1 $\times 10^{-7}\;and 1.1 \times 10^{-6}$ M, respectively. MR-387Al also strongly inhibited aminopeptidase B of human myelogenous leukemia K-562 cell with $IC_50$ of 0.35 $\mu$ M. Inhibitions of aminopeptidase N activity by ARPA-bearing inhibitors of various peptide chain lengths also have been studied. $IC_ 50$ values of AHPA-Val (bestatin), ARPA-Val-Pro (MR-387Al) and ARPA-Val-Pro-Leu (MR-387C) compared against porcine kidney aminopeptidase N were 20.1, 0.60 and 0.08 $\mu$ M, respectively. These results support that a multiple interaction between the $S_1\to S'_3$ sites of aminopeptidase N and the $P_1\to P'_3$ of the inhibitor plays a crucial role in stabilizing strongly the enzyme-inhibitor complex.

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OECD/NEA BENCHMARK FOR UNCERTAINTY ANALYSIS IN MODELING (UAM) FOR LWRS - SUMMARY AND DISCUSSION OF NEUTRONICS CASES (PHASE I)

  • Bratton, Ryan N.;Avramova, M.;Ivanov, K.
    • Nuclear Engineering and Technology
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    • v.46 no.3
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    • pp.313-342
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    • 2014
  • A Nuclear Energy Agency (NEA), Organization for Economic Co-operation and Development (OECD) benchmark for Uncertainty Analysis in Modeling (UAM) is defined in order to facilitate the development and validation of available uncertainty analysis and sensitivity analysis methods for best-estimate Light water Reactor (LWR) design and safety calculations. The benchmark has been named the OECD/NEA UAM-LWR benchmark, and has been divided into three phases each of which focuses on a different portion of the uncertainty propagation in LWR multi-physics and multi-scale analysis. Several different reactor cases are modeled at various phases of a reactor calculation. This paper discusses Phase I, known as the "Neutronics Phase", which is devoted mostly to the propagation of nuclear data (cross-section) uncertainty throughout steady-state stand-alone neutronics core calculations. Three reactor systems (for which design, operation and measured data are available) are rigorously studied in this benchmark: Peach Bottom Unit 2 BWR, Three Mile Island Unit 1 PWR, and VVER-1000 Kozloduy-6/Kalinin-3. Additional measured data is analyzed such as the KRITZ LEU criticality experiments and the SNEAK-7A and 7B experiments of the Karlsruhe Fast Critical Facility. Analyzed results include the top five neutron-nuclide reactions, which contribute the most to the prediction uncertainty in keff, as well as the uncertainty in key parameters of neutronics analysis such as microscopic and macroscopic cross-sections, six-group decay constants, assembly discontinuity factors, and axial and radial core power distributions. Conclusions are drawn regarding where further studies should be done to reduce uncertainties in key nuclide reaction uncertainties (i.e.: $^{238}U$ radiative capture and inelastic scattering (n, n') as well as the average number of neutrons released per fission event of $^{239}Pu$).

Purification and Characterization of Extracellular Aspartic Proteinase of Candida albicans

  • Na, Byoung-Kuk;Lee, Seong-Il;Kim, Sin-Ok;Park, Young-Kil;Bai, Gill-Han;Kim, Sang-Jae;Song, Chul-Yong
    • Journal of Microbiology
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    • v.35 no.2
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    • pp.109-116
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    • 1997
  • An extracellular proteinase of Candida albicans was purified by a combination of 0~75% ammonium sulfate precipitation, DEAE Sepharose Fast Flow ion exchange chromatography, and Sephacryl S-200 HR molecular sieve chromatography. Its mlecular weight was approximately 41 kDa on SDS-PAGE and isoelectric point was 4.4. The enzyme was inhibited by pepstain A. Optimum enzyme activity ranged from pH 2.0 to 3.5 with its maximum at pH 2.5 and a temperature of 45$^{\circ}C$. The addition of divalent cations, $Ca^{2+}$, Zn$^{2+}$ and $Mg^{2+}$, resulted in no significant inhibition of enzymatic activity. However, some inhibitory effects were observed by Fe$^{2+}$, Ag$^{2+}$ and Cu$^{2+}$. With BSA as substrate, an apparent $K_m$ was determined to be 7$\times$10$^{-7}$ M and $K_i$, using pepstatin A as an inhibitor, was 8.05$\times$10$^{-8}$ M. N-terminal amino acid sequence was QAVPVTLXNEQ. Degradation of BSA and fibronectin was shown but not collagen, hemoglobin, immunoglobulin G, or lysozyme. The enzyme preferred peptides with Glu and Leu at the P$_1$ position, but the enzyme activity was highly reduced when the P$_2$ position was phe or pro. This enzyme showed antigenicity against sera of patients with candidiasis.

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ermK Leader Peptide : Amino Acid Sequence Critical for Induction by Erythromycin

  • Kwon, Ae-Ran;Min, Yu-Hong;Yoon, Eun-Jeong;Kim, Jung-A;Shim, Mi-Ja;Choi, Eung-Chil
    • Archives of Pharmacal Research
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    • v.29 no.12
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    • pp.1154-1157
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    • 2006
  • The ermK gene from Bacillus lichenformis encodes an inducible rRNA methylase that confers resistance to the macrolide-lincosamide-streptogramin B antibiotics. The ermK mRNA leader sequence has a total length of 357 nucleotides and encodes a 14-amino acid leader peptide together with its ribosome binding site. The secondary structure of ermK leader mRNA and a leader peptide sequence have been reported as the elements that control expression. In this study, the contribution of specific leader peptide amino acid residues to induction of ermK was studied using the PCR-based megaprimer mutation method. ermK methylases with altered leader peptide codons were translationally fused to E. coli ${\beta}-galactosidase$ reporter gene. The deletion of the codons for Thr-2 through Ser-4 reduced inducibility by erythromycin, whereas that for Thr-2 and His-3 was not. The replacement of the individual codons for Ser-4, Met-5 and Arg-6 with termination codon led to loss of inducibility, but stop mutation of codon Phe-9 restored inducibility by erythromycin. Collectively, these findings suggest that the codons for residue 4, 5 and 6 comprise the critical region for induction. The stop mutation at Leu-7 expressed constitutively ermK gene. Thus, ribosome stalling at codon 7 appears to be important for ermK induction.

Purification and Properties of Osteopontin from Bovine Milk (우유로부터 Osteopontin의 분리.정제 및 특성에 관한 연구)

  • Choi, K.W.;Kim, D.W.;Lee, S.W.
    • Journal of Animal Science and Technology
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    • v.45 no.3
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    • pp.491-498
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    • 2003
  • The purpose of this study is to observe purification and properties of osteopontin(OPN) from bovine milk. The purification of osteopontin from bovine milk was performed by using ion-exchange and hydrophobic chromatography. SDS-PAGE analysis revealed that the protein migrated at Mw. 60,000. NH2-terminal sequence analysis of the first seven amio acids revealed the protein to be identical to that previously reported for bovine OPN. 35-wk-old chickens, including 3 Single Comb White Leghorn (SCWL), were used to produce egg yolk antibody(IgY) against OPNas a antigen. However, the anti-OPN antibody activities determined by ELISA. Immunological assy of OPN in milk was performed using radial immunodiffusion test based on the standard curve of pure OPN. The radial precipitation lines of four different milk samples indicated that the concentrations of OPN in the milk samples were within the range of 31.7 to 39.7${\mu}g$/ml. On inhibition with OPN on precipitation of calcium phosphate, OPN was slightly higher than casein phosphopeptide(CPP) and poly-glutamic acid.

Distal Chevron Osteotomy for the Treatment of Moderate and Severe Hallux Valgus (증등도 및 중증 무지 외반증 치료에서 원위부 갈매기형 절골술의 적용)

  • Yoon, Jun-O;Lee, Ho-Seung;Leu, Suk-Woo;Lee, Ki-Won;Oh, Se-Kwan
    • Journal of Korean Foot and Ankle Society
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    • v.7 no.2
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    • pp.166-173
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    • 2003
  • Purpose: We assessed the treatment result of the distal chevron osteotomy in the patients with moderate to severe hallux valgus. Materials and Methods: In a total of 28 cases of hallux valgus in 20 patients, underwent distal chevron osteotomy between July 1999 and February 2001, were enrolled in this study. 21 cases were moderate and 7 cases were severe. The preoperative average hallux valgus angle and 1st-2nd intermetatarsal angle of the two groups were $31.5^{\circ}$, $15.8^{\circ}$ in moderate cases and $44.1^{\circ}$, $17.3^{\circ}$ in severe cases, respectively. Radiologic evaluation was done preoperatively, postoperatively and on the final follow-up visit using weight-bearing radiographic imaging to determine the hallux valgus angle and 1st-2nd intermetatarsal angle. Results: Radiographic evaluation revealed hallux valgus angle and 1st-2nd intermetatarsal angle in moderate cases to be $13.0^{\circ}$, $11.3^{\circ}$ (postoperatively and in severe cases $15.6^{\circ}$, $10.9^{\circ}C$, postoperatively. On final follow up, the results were $14.5^{\circ}$, $11.6^{\circ}$ in moderate cases and $18.3^{\circ}$, $11.9^{\circ}$ in severe cases, respectively. Conclusion: Distal chevron osteotomy can be usefully applied to the treatment of moderate to severe hallux valgus.

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In-silico and structure-based assessment to evaluate pathogenicity of missense mutations associated with non-small cell lung cancer identified in the Eph-ephrin class of proteins

  • Shubhashish Chakraborty;Reshita Baruah;Neha Mishra;Ashok K Varma
    • Genomics & Informatics
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    • v.21 no.3
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    • pp.30.1-30.13
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    • 2023
  • Ephs belong to the largest family of receptor tyrosine kinase and are highly conserved both sequentially and structurally. The structural organization of Eph is similar to other receptor tyrosine kinases; constituting the extracellular ligand binding domain, a fibronectin domain followed by intracellular juxtamembrane kinase, and SAM domain. Eph binds to respective ephrin ligand, through the ligand binding domain and forms a tetrameric complex to activate the kinase domain. Eph-ephrin regulates many downstream pathways that lead to physiological events such as cell migration, proliferation, and growth. Therefore, considering the importance of Eph-ephrin class of protein in tumorigenesis, 7,620 clinically reported missense mutations belonging to the class of variables of unknown significance were retrieved from cBioPortal and evaluated for pathogenicity. Thirty-two mutations predicted to be pathogenic using SIFT, Polyphen-2, PROVEAN, SNPs&GO, PMut, iSTABLE, and PremPS in-silico tools were found located either in critical functional regions or encompassing interactions at the binding interface of Eph-ephrin. However, seven were reported in nonsmall cell lung cancer (NSCLC). Considering the relevance of receptor tyrosine kinases and Eph in NSCLC, these seven mutations were assessed for change in the folding pattern using molecular dynamic simulation. Structural alterations, stability, flexibility, compactness, and solvent-exposed area was observed in EphA3 Trp790Cys, EphA7 Leu749Phe, EphB1 Gly685Cys, EphB4 Val748Ala, and Ephrin A2 Trp112Cys. Hence, it can be concluded that the evaluated mutations have potential to alter the folding pattern and thus can be further validated by in-vitro, structural and in-vivo studies for clinical management.

Study on the Processing and Compositions of Salted and Dried Mullet Roe (영암산 염건 숭어알의 가공과 조성에 관한 연구)

  • Joe, Sang-June;Rhee, Chong-Ouk;Kim, Dong-Youn
    • Korean Journal of Food Science and Technology
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    • v.21 no.2
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    • pp.242-251
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    • 1989
  • The salted-dried mullet(Mugil japonicus) roe is a kind of traditional food particulary in the area of Young-am gun, Chunnam province. This study was conducted to conform the scientific processing conditions and to evaluate the nutritional quality and changes of major components during storage times. The manufacturing method was that the fresh roe was salted for about 20 hours for the preparation of salted-dried roe, washed by clean waters, drained, shaped a flat piece with 1.2cm thickness by pressing, and spreaded sesame oils on the surface of the salted roe periodically during wind drying for 20 days. The dried roe was blanched in heated water$(80^{\circ}C/3min)$ and packaged the dried product for storages. The fractional compositions of free lipid of wind dried roe were 40% of neutral lipids, 12% of glycolipids and 9% of phospholipids and those of bound lipids were 13% of neutral lipids. 10% of glycolipids and 13% of phospholipids respectively. The major fatty acids of the roe were $C_{16:0}$, $C_{18:0}$, $C_{18:1}$, $C_{18:2}$ and $C_{20:0}$ which was consisted of free and bound lipids in wind drying method during processing and storages. Total amino acids were 99.87g/100g and major amino acids were Glu, Pro, Leu, Lys and CySH and the protein score was average 155% and the chemical score was average 109%. Free amino acids was 1,376mg% that had 50.61% of Pro and the major kinds of those were Tyr and CySH.

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