• Title/Summary/Keyword: amino acid ester

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Novel Glycolipoproteins from Ginseng

  • Pyo, Mi-Kyung;Choi, Sun-Hye;Hwang, Sung-Hee;Shin, Tae-Joon;Lee, Byung-Hwan;Lee, Sang-Mok;Lim, Yoong-Ho;Kim, Dong-Hyun;Nah, Seung-Yeol
    • Journal of Ginseng Research
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    • v.35 no.1
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    • pp.92-103
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    • 2011
  • Ginseng has been used as a general tonic agent to invigorate human body. In the present study, we isolated novel glycolipoproteins from ginseng that activate $Ca^{2+}$-activated $Cl^-$ channel (CaCC) in Xenopus oocytes and transiently increase intracellular free $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) in mouse Ehrlich ascites tumor cells. We named the active ingredients as gintonin. Gintonin exists in at least six different forms. The native molecular weight of gintonin is about 67 kDa but its apparent molecular weight is about 13 kDa, indicating that gintonin might be a pentamer. Gintonin is rich in hydrophobic amino acids. Its main carbohydrates are glucose and glucosamine. Its lipid components are linoleic, palmitic, oleic, and stearic acids. Gintonin actions were blocked by U73122, a phospholipase C inhibitor, 2-aminoethxydiphenyl borate, an inositol 1,4,5-trisphosphate receptor antagonist, or bis (o-aminophenoxy) ethane-N,N,N0,N0-tetracetic acid acetoxymethyl ester, a membrane permeable $Ca^{2+}$ chelator. In the present study, we for the first time isolated novel gintonin and showed the signaling pathways on gintonin-mediated CaCC activations and transient increase of $[Ca^{2+}]_i$. Since $[Ca^{2+}]_i$ as a second messenger plays a pivotal role in the regulation of diverse $Ca^{2+}$-dependent intracellular signal pathways, gintonin-mediated regulations of $[Ca^{2+}]_i$ might contribute to biological actions of ginseng.

Biochemical Characterization of a Novel Thermostable Esterase from the Metagenome of Dokdo Islets Marine Sediment (독도 심해토 메타게놈 유래 신규 내열성 에스테라아제의 생화학적 특성규명)

  • Lee, Chang-Muk;Seo, Sohyeon;Kim, Su-Yeon;Song, Jaeeun;Sim, Joon-Soo;Hahn, Bum-Soo;Kim, Dong-Hern;Yoon, Sang-Hong
    • Microbiology and Biotechnology Letters
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    • v.45 no.1
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    • pp.63-70
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    • 2017
  • A functional screen of 60,672 fosmid metagenomic clones amplified from marine sediment obtained from the Dokdo islets in Korea identified the gene EstES1, whose product, EstES1, displayed lipolytic properties on tributyrin-supplemented media. EstES1 is a 576 amino acid protein with a predicted molecular weight of 59.4 kDa including 37 N-terminal leader amino acids. EstES1 exhibited the highest sequence similarity (44%) to a carboxylesterase found in Haliangium ochraceum DSM14365. Phylogenetic analysis indicated that EstES1 belongs to a currently uncharacterized family of lipases. Within the conserved domain, EstES1 retains the catalytic triad that consists of the consensus penta-peptide motif, GESAG. EstES1 demonstrated a broad substrate specificity toward the long acyl group of ethyl esters (C2-C12), and its optimal activity was recorded toward p-Nitrophenyl butyrate (C4) at pH 9.0 and $40^{\circ}C$ (specific activity of 255.4 U/mg). The enzyme remained stable in the ranges of $60-65^{\circ}C$ and pH 9.0-10.5 and in the presence of methanol, ethanol, isopropanol, and dimethyl sulfoxide. Therefore, EstES1 has potential for use in industrial applications involving high temperature, organic solvents, and/or alkaline conditions.

ATHEROSCLEROSIS, CHOLESTEROL AND EGG - REVIEW -

  • Paik, I.K.;Blair, R.
    • Asian-Australasian Journal of Animal Sciences
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    • v.9 no.1
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    • pp.1-25
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    • 1996
  • The pathogenesis of atherosclerosis can not be summarized as a single process. Lipid infiltration hypothesis and endothelial injury hypothesis have been proposed and investigated. Recent developments show that there are many points of potential interactions between them and that they can actually be regarded as two phases of a single, unifying hypothesis. Among the many risk factors of atherosclerosis, plasma homocysteine and lipoprotein(a) draw a considerable interest because they are independent indicators of atherogenicity. Triglyceride (TG)-rich lipoproteins (chylomicron and VLDL) are not considered to be atherogenic but they are related to the metabolism of HDL cholesterol and indirectly related to coronary heart disease (CHD). LDL can of itself be atherogenic but the oxidative products of this lipoprotein are more detrimental. HDL cholesterol has been considered to be a favorable cholesterol. The so-called 'causalist view' claims that HDL traps excess cholesterol from cellular membranes and transfers it to TG-rich lipoproteins that are subsequently removed by hepatic receptors. In the so-called 'noncausalist view', HDL does not interfere directly with cholesterol deposition in the arterial wall but instead reflects he metabolism of TG-rich lipoproteins and their conversion to atherogenic remnants. Approximately 70-80% of the human population shows an effective feedback control mechanism in cholesterol homeostasis. Type of dietary fat has a significant effect on the lipoprotein cholesterol metabolism and atherosclerosis. Generally, saturated fatty acids elevate and PUFA lower serum cholesterol, whereas MUFA have no specific effect. EPA and DHA inhibit the synthesis of TG, VLDL and LDL, and may have favourable effects on some of the risk factors. Phospholipids, particularly lecithin, have an antiatherosclerotic effect. Essential phospholipids (EPL) may enhance the formation of polyunsaturated cholesteryl ester (CE) which is less sclerotic and more easily dispersed via enhanced hydrolysis of CE in the arterial wall. Also, neutral fecal steroid elimination may be enhanced and cholesterol absorption reduced following EPL treatment. Antioxidants protect lipoproteins from oxidation, and cells from the injury of toxic, oxidized LDL. The rationale for lowering of serum cholesterol is the strong association between elevation of plasma or serum cholesterol and CHD. Cholesterol-lowing, especially LDL cholesterol, to the target level could be achieved using diet and combination of drug therapy. Information on the link between cholesterol and CHD has decreased egg consumption by 16-25%. Some clinical studies have indicated that dietary cholesterol and egg have a significant hypercholesterolemic effect, while others have indicated no effect. These studies differed in the use of purified cholesterol or cholesterol in eggs, in the range of baseline and challenge cholesterol levels, in the quality and quantity of concomitant dietary fat, in the study population demographics and initial serum cholesterol levels, and clinical settings. Cholesterol content of eggs varies to a certain extent depending on the age, breed and diet of hens. However, egg yolk cholesterol level is very resistant to change because of the particular mechanism involved in yolk formation. Egg yolk contains a factor of factors responsible for accelerated cholesterol metabolism and excretion compared with crystalline cholesterol. One of these factors could be egg lecithin. Egg lecithin may not be as effective as soybean lecithin in lowering serum cholesterol level due probably to the differences of fatty acid composition. However, egg lecithin may have positive effects in hypercholesterolemia by increasing serum HDL level and excretion of fecal cholesterol. The association of serum cholesterol with egg consumption has been widely studied. When the basal or control diet contained little or no cholesterol, consumption of 1 or 2 eggs daily increased the concentration of plasma cholesterol, whereas that of the normolipemic persons on a normal diet was not significantly influenced by consuming 2 to 3 eggs daily. At higher levels of egg consumption, the concentration of HDL tends to increase as well as LDL. There exist hyper-and hypo-responders to dietary (egg) cholesterol. Identifying individuals in both categories would be useful from the point of view of nutrition guidelines. Dietary modification of fatty acid composition has been pursued as a viable method of modifying fat composition of eggs and adding value to eggs. In many cases beneficial effects of PUFA enriched eggs have been demonstrated. Generally, consumption of n-3 fatty acids enriched eggs lowered the concentration of plasma TG and total cholesterol compared to the consumption of regular eggs. Due to the highly oxidative nature of PUFA, stability of this fat is essential. The implication of hepatic lipid accumulation which was observed in hens fed on fish oils should be explored. Nutritional manipulations, such as supplementation with iodine, inhibitors of cholesterol biosynthesis, garlic products, amino acids and high fibre ingredients, have met a limited success in lowering egg cholesterol.

Synthesis and Biodistribution of Flumazenil Derivative [F-18](3-(2-Fluoro) flumazenil for Imaging Benzodiazepine Receptor (벤조디아제핀 수용체 영상용 양전자 방출 핵종 표지 플루마제닐 유도체 [F-18](3-(2-Fluoro)flumazenil의 합성과 생체 내 분포)

  • Hong, Sung-Hyun;Jeong, Jae-Min;Chang, Young-Soo;Lee, Dong-Soo;Chung, June-Key;Cho, Jung-Hyuck;Lee, Sook-Ja;Kang, Sam-Sik;Lee, Myung-Chul
    • The Korean Journal of Nuclear Medicine
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    • v.33 no.6
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    • pp.527-536
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    • 1999
  • Purpose: Radiotracers that bind to the central benzodiazepine receptor are useful for the investigation of various neurological and psychiatric diseases. [C-11]Flumazenil, a benzodiazepine antagonist, is the most widely used radioligand for central benzodiazepine receptor imaging by PET. We synthesized 3-(2-[F-18]fluoro)flumazenil, a new fluorine-18 ($t_{1/2}$= 110 min) labeled analogue of benzodiazepine receptor imaging agent, and evaluated in vivo for biodistribution in mice. Materials and Methods: Flumazenil (Ro 15-1788) was synthesized by a modification of the reported method. Precursor of 3-(2-[F-18]fluoro)flumazenil, the tosylated flumazenil derivative was prepared by the tosylation of the ethyl ester by ditosylethane. [F-18] labeling of tosyl substitued flumazenil precursor was performed by adding F-18 ion at $85^{\circ}C$ in the hot ceil for 20 min. The reaction mixture was trapped by C18 cartridge, washed with 10% ethanol, and eluted by 40% ethanol. Bidistribution in mice was determined after intravenous injection. Results: The total chemical yield of tosylated flumazenil derivative was ${\sim}40%$. The efficiency of labeling 3-(2-[F-18]fluoro)flumazenil was 66% with a total synthesis time of 50 min. Brain uptakes of 3-(2-[F-18]fluoro)flumazenil at 10, 30, 60 min after injection, were $2.5{\pm}0.37,\;2.2{\pm}0.26,\;2.1{\pm}0.11$ and blood activities were $3.7{\pm}0.43,\;3.3{\pm}0.07,\;3.3{\pm}0.09%ID/g$, respectively. Conclusion: We synthesized a tosylated flumazenil derivative which was successfully labeled with no-carrier-added F-18 by nucleophilic substitution.

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A novel cold-active lipase from Psychrobacter sp. ArcL13: gene identification, expression in E. coli, refolding, and characterization (새로운 Psychrobacter sp. ArcL13 유래 저온활성 지질분해효소 : 유전자 분리동정, 대장균에서의 발현, refolding 및 특성 연구)

  • Koo, Bon-Hun;Moon, Byung-Hern;Shin, Jong-Suh;Yim, Joung-Han
    • Korean Journal of Microbiology
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    • v.52 no.2
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    • pp.192-201
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    • 2016
  • Recently, Psychrobacter sp. ArcL13 strain showing the extracellular lipase activity was isolated from the Chuckchi Sea of the Arctic Ocean. However, due to the low expression levels of the enzyme in the natural strain, the production of recombinant lipase is crucial for various applications. Identification of the gene for the enzyme is prerequisite for the production of the recombinant protein. Therefore, in the present study, a novel lipase gene (ArcL13-Lip) was isolated from Psychrobacter sp. ArcL13 strain by gene prospecting using PCR, and its complete nucleotide sequence was determined. Sequence analysis showed that ArcL13-Lip has high amino acid sequence similarity to lipases from bacteria of some Psychrobacter genus (84-90%) despite low nucleotide sequence similarity. The lipase gene was cloned into the bacterial expression plasmid and expressed in E. coli. SDS-PAGE analysis of the cells showed that ArcL13-Lip was expressed as inclusion bodies with a molecular mass of about 35 kDa. Refolding was achieved by diluting the unfolded protein into refolding buffers containing various additives, and the highest refolding efficiency was seen in the glucose-containing buffer. Refolded ArcL13-Lip showed high hydrolytic activity toward p-nitrophenyl caprylate and p-nitrophenyl decanoate among different p-nitrophenyl esters. Recombinant ArcL13-Lip displayed maximal activity at $40^{\circ}C$ and pH 8.0 with p-nitrophenyl caprylate as a substrate. Activity assays performed at various temperatures showed that ArcL13-Lip is a cold-active lipase with about 40% and 73% of enzymatic activity at $10^{\circ}C$ and $20^{\circ}C$, respectively, compared to its maximal activity at $40^{\circ}C$.