• 자연과학논문집
• /
• 제9권1호
• /
• pp.25-29
• /
• 1997

Glucocorticoid에 의한 항염증 작용의 second messenger로 생각되어지는 annexin superfamily중 하나인 37 kDa의 단백질, lipocortin-1의 작용기작을 이해할 목적으로 in vivo에서 protein-protein interaction을 인식하는 yeast-based genetic assay인 yeast two assay를 통하여 lipocortin-1과 결합하는 단백질 유전자를 분리하여 조사하였다. 이 방법으로 실험을 수행한 결과 분리된 유전자가 human serine proteinase 유전자와 homology가 있는 것으로 밝혀졌다.

• Journal of Microbiology
• /
• 제35권2호
• /
• pp.123-126
• /
• 1997

The aim of the present study was to optimize culture condition for the expression of lipocortin-1$_{1-185}$ in a recombinant of Escherichia coli using batch system. Plasmid (pHT22) carrying lipocortin-1$_{1-185}$ gene was well maintained in the recombinant with the addition of amplicillin as a selection pressures. Optimum temperature was 28$^{\circ}C$ for seed culture and 4$0^{\circ}C$ for main culture and the optimum pH was 7.0. The production of Lipocortin-1$_{1-185}$ was closely associated with cell growth and related to plasmid amplification.

• The Korean Journal of Physiology and Pharmacology
• /
• 제1권1호
• /
• pp.91-96
• /
• 1997

Lipocortin 1 has been proposed as a putative mediator of anti-inflammatory actions of glucocorticoids. We investigated the role of lipocortin 1 in the effect of dexamethasone using rat splenic leukocytes. Concanavalin A(ConA; 1 ${\mu}g/ml$) increased the leukocyte proliferation and nitric oxide(NO) generation, which were measured as $[^3H]-thymidine$ uptake by the cells and nitrite accumulation in the culture media, respectively. Dexamethasone suppressed ConA-induced cell proliferation, in a concentration-dependent manner with $EC_{50}$ around 50nM. The addition of anti-lipocortin l(Anti-LCl) reversed dexamethasone effects: 0.24, 1.2, 6 ${\mu}g/ml$ of Anti-LC1 reversed dexamethasone(50 nM)-induced suppression of thymidine uptake by $9{\pm}3%$, $16{\pm}3%$, $36{\pm}5%$, respectively; 0.24, 1.2, and 6 ${\mu}g/ml$ of Anti-LCI reversed dexa-methasone-induced decrease of nitrite concentration by $49{\pm}16%$, $61{\pm}20%$, $77{\pm}19%$, respectively. The present data indicate that lipocortin 1 mediates, at least in part, glucocorticoids-induced suppression of leukocyte proliferation and blockade of NO generation.

• Journal of Microbiology and Biotechnology
• /
• 제3권3호
• /
• pp.168-173
• /
• 1993

The expression kinetics of human lipocortin I (LCI), a potential anti-inflammatory agent, was studied in the shake-flask and fermenter cultures of Saccharomyces cerevisiae carrying a galactose-inducible expression system. The cell growth, expression level of LCI, and the plasmid stability were investigted under various galactose induction conditions. The expression of LCI was repressed by the presence of a very small amount of dextrose in the culture medium, but it was induced by galactose after dextrose became completely depleted. The optimal ratio of dextrose to galactose for lipocortin I production was found to be 1.0 (10 g/l dextrose and 10 g/l galactose). With optimal D/G ratio of 1.0 and the addition of galactose prior to dextrose depletion, LCI of about 100~130 mg/l was produced. LCI at a concentration of 174 mg/l was porduced in the fed-batch culture, which was nearly a twice as much of that produced in the batch culture. The plasmid stability was very high in all culture cases, and thus was considered to be not an important parameter in the expression of LCI.

• BMB Reports
• /
• 제31권2호
• /
• pp.117-122
• /
• 1998

Changes of lipocortin 1 (LC1) in the brain induced by immobilization stress were investigated in rats. Rats were immobilized for 0,1,2,3,4, and 5 h, and the brain slices were immunostained with anti-human LC1 antibodl (anti-LC1). Immunoreactivity of LCI (iLC1) was most prominent in neuronal cell bodies and processes of hippocampal CA regions and dentate gyrus. At rest without stress, most of the LC1 in the neuron located in the cytoplasm with the nuclei exhibiting relatively scarce immunoreactivity. Immobilization stress changed this intracellular distribution of LC1 by increasing nuclear LC1. The change was apparent in 1 h and reached the peak by 3 h. However, by 5 h of immobilization, the distribution pattern returned to that of the resting state. This transient nuclear translocation of LC1 was most prominent in $CA_1$ pyramidal neurons, and was not observed in areas other than the hippocampus. Adrenalectomy abolished this transient translocation of LC1. The roles of hippocampal LC1 as a mediator of glucocorticoid feedback signal and/or as an intracellar stress signaling protein could be suggested.

• Journal of Microbiology and Biotechnology
• /
• 제5권4호
• /
• pp.224-228
• /
• 1995

Fed-batch fermentations were conducted to produce human lipocortin-I (LC1), a potential anti-inflammatory agent, from recombinant Sacchromyces cerevisiae carrying a galactose-inducible expression system. The cell growth, expression level of LC1, and the plasmid stability were investigated under various LC1 induction modes performed by three different galactose feeding strategies. Galactoe was fed to induce the expression of LCl from the beginning (initial induction) of culture or when the cell concentration reached 120 OD (mid-phase induction) or 300 OD (late induction). Among the three galactose-induction modes tested, the initial induction mode yielded the best result with respect to a final expression level of LC1. Fedbatch fermentation with initial induction mode produced LC1 at a conentration of 220 mg/l, which corresponded to 1.38- and 1.53-fold increases over those produced by mid-phase and late induction modes.

• Journal of Microbiology and Biotechnology
• /
• 제4권4호
• /
• pp.237-244
• /
• 1994

The vector systems for the expression and secretion of human lipocortin-l (LC1) from Saccharomyces cerevisiae were constructed with GAL10 promoter and the prepro leader sequence of mating factor-$\alpha$1. They were further constructed to contain three different transcription terminators; GAL7 terminator, LCl terminator and a fused form of these two terminators. The expression and secretion levels of LCl were compared to investigate the effect of transcription terminators on the LCl gene expression. For the expression cassettes employing the GAL7 terminator or the terminator of fused form, the expression levels of LCl were measured by scanning the immunoreactive LCl protein bands, and were found to be 0.27 g/l and 0.32 g/l, respectively. The highest expression level of 0.54 g/l was obtained with the expression vector containing the LCl transcription terminator. In all expression cassettes, the majority of LCl proteins expressed were retained intracellularly, indicating a low secretion efficiency of about 5%. The high expression level of LCl was explained by the great content and stability of LCl mRNA transcribed from the LCl terminator-employing vector. The results of this study demonstrate that the LCl transcription terminator functions for the expression of LCl in S. cerevisiae better than the GAL7 terminator.

• 한국동물학회:학술대회논문집
• /
• 한국동물학회 1995년도 한국생물과학협회 학술발표대회
• /
• pp.355.1-355
• /
• 1995

No Abstract, See Full Text

• BMB Reports
• /
• 제32권1호
• /
• pp.28-32
• /
• 1999

Annexin I (also called lipocortin 1), a 37-kDa member of the annexin family of proteins, has been implicated in the mitogenic signal transduction by epidermal growth factor (EGF). Annexin I is phosphorylated by the EGF signal, however, the role of annexin I in the EGF signal transduction is still unknown. To transduce extracellular signals into the intracellular targets, selective translocation of the signaling molecules to their targets would be necessary. In this study, we examined the subcellular locations of annexin I during EGF signal transduction. Treatment of A549 cells with EGF resulted in the translocation of cytoplasmic annexin I to the nucleus and perinuclear region as determined by Western blot and immunofluorescent staining. The nuclear translocation of annexin I was inhibited by tyrphostin AG 1478 and genistein, the inhibitors of EGF receptor kinase and downstream tyrosine kineses, respectively. Pretreatment of cells with cyclohexamide did not inhibit the nuclear translocation. The results suggest that nuclear translocation of annexin I is controlled by a series of kinase dependent events in the EGF receptor signaling pathway and may be important in tranducing the signals by EGF.