共查询到20条相似文献,搜索用时 255 毫秒
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14-3-3蛋白家族是由多个高度保守的成员构成的调节性蛋白质家族,它们主要以磷酸化的形式与伴侣蛋白相互作用,并能够以多种方式来影响靶蛋白。通过构建14-3-3蛋白原核表达载体,纯化重组蛋白获得14-3-3蛋白抗体。为了验证14-3-3蛋白基因在耐铝中的作用,构建14-3-3酵母表达载体,得到14-3-3过表达酵母菌株。在5mmol/L铝浓度下,转基因酵母比对照酵母长势好,这表明14-3-3蛋白通过促进生长赋予酵母对铝胁迫的耐受性。 相似文献
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In this study, the interaction between rice 14-3-3 protein and 1-aminocyclopropane carboxylic acid synthase (ACS) was observed
in yeast cells using yeast two-hybrid assays. Given the fact that 14-3-3 proteins generally bind to their target proteins
in a phosphorylation-dependent manner, a hypothesis regarding the regulatory role of 14-3-3 proteins in the activation of
ACS is proposed in which 14-3-3 proteins may bind to the phosphorylated C-terminal tails of ACSs and help them to escape from
their fated degradation when ethylene biosynthesis is needed. It is reasonable to believe that 14-3-3 protein may play an
important role in regulating ACS activity.
Published in Russian in Biokhimiya, 2007, Vol. 72, No. 9, pp. 1231–1237. 相似文献
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Under drought stress, ABA promotes stomatal closure to prevent water loss. Although protein phosphorylation plays an important role in ABA signaling, little is known about these processes at the biochemical level. In this study, we searched for substrates of protein kinases in ABA signaling through the binding of a 14-3-3 protein to phosphorylated proteins using Vicia guard cell protoplasts. ABA induced binding of a 14-3-3 protein to proteins with molecular masses of 61, 43 and 39 kDa, with the most remarkable signal for the 61 kDa protein. The ABA-induced binding to the 61 kDa protein occurred only in guard cells, and reached a maximum within 3 min at 1 microM ABA. The 61 kDa protein localized in the cytosol. ABA induced the binding of endogenous vf14-3-3a to the 61 kDa protein in guard cells. Autophosphorylation of ABA-activated protein kinase (AAPK), which mediates anion channel activation, and ABA-induced phosphorylation of the 61 kDa protein showed similar time courses and similar sensitivities to the protein kinase inhibitor K-252a. AAPK elicits the binding of the 14-3-3 protein to the 61 kDa protein in vitro when AAPK in guard cells was activated by ABA. The phosphorylation of the 61 kDa protein by ABA was not affected by the NADPH oxidase inhibitor, H(2)O(2), W-7 or EGTA. From these results, we conclude that the 61 kDa protein may be a substrate for AAPK and that the 61 kDa protein is located upstream of H(2)O(2) and Ca(2+), or on Ca(2+)-independent signaling pathways in guard cells. 相似文献
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Kim D Won J Shin DW Kang J Kim YJ Choi SY Hwang MK Jeong BW Kim GS Joe CO Chung SH Song WJ 《Biochemical and biophysical research communications》2004,323(2):499-504
Dual-specificity tyrosine(Y) regulated kinase 1A (DYRK1A) is a serine/threonine protein kinase implicated in mental retardation resulting from Down syndrome. In this study, we carried out yeast two-hybrid screening to find proteins regulating DYRK1A kinase activity. We identified 14-3-3 as a Dyrk1A interacting protein, which is consistent with the previous finding of the interaction between the yeast orthologues Yak1p and Bmh1/2p. We showed the interaction between Dyrk1A and 14-3-3 in vitro and in vivo. The binding required the N-terminus of Dyrk1A and was independent of the Dyrk1A phosphorylation status. Functionally, 14-3-3 binding increased Dyrk1A kinase activity in a dose dependent manner in vitro. In vivo, a small peptide inhibiting 14-3-3 binding, sc138, decreased Dyrk1A kinase activity in COS7. In summary, these results suggest that DYRK1A kinase activity could be regulated by the interaction of 14-3-3. 相似文献
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Molecular evolution of the 14-3-3 protein family 总被引:9,自引:0,他引:9
Members of the highly conserved and ubiquitous 14-3-3 protein family modulate a wide variety of cellular processes. To determine
the evolutionary relationships among specific 14-3-3 proteins in different plant, animal, and fungal species and to initiate
a predictive analysis of isoform-specific differences in light of the latest functional and structural studies of 14-3-3,
multiple alignments were constructed from forty-six 14-3-3 sequences retrieved from the GenBank and SwissProt databases and
a newly identified second 14-3-3 gene fromCaenorhabditis elegans. The alignment revealed five highly conserved sequence blocks. Blocks 2–5 correlate well with the alpha helices 3, 5, 7,
and 9 which form the proposed internal binding domain in the three-dimensional structure model of the functioning dimer. Amino
acid differences within the functional and structural domains of plant and animal 14-3-3 proteins were identified which may
account for functional diversity amongst isoforms. Protein phylogenic trees were constructed using both the maximum parsimony
and neighbor joining methods of the PHYLIP(3.5c) package; 14-3-3 proteins fromEntamoeba histolytica, an amitochondrial protozoa, were employed as an outgroup in our analysis. Epsilon isoforms from the animal lineage form
a distinct grouping in both trees, which suggests an early divergence from the other animal isoforms. Epsilons were found
to be more similar to yeast and plant isoforms than other animal isoforms at numerous amino acid positions, and thus epsilon
may have retained functional characteristics of the ancestral protein. The known invertebrate proteins group with the nonepsilon
mammalian isoforms. Most of the current 14-3-3 isoform diversity probably arose through independent duplication events after
the divergence of the major eukaryotic kingdoms. Divergence of the seven mammalian isoforms beta, zeta, gamma, eta, epsilon,
tau, and sigma (stratifin/ HME1) occurred before the divergence of mammalian and perhaps before the divergence of vertebrate
species. A possible ancestral 14-3-3 sequence is proposed.
Correspondence to: D.C. Shakes 相似文献
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The 14-3-3 protein family interacts with more than 2000 different proteins in mammals, as a result of its specific phospho-serine/phospho-threonine binding activity. Seven paralogs are strictly conserved in mammalian species. Here, we show that during adipogenic differentiation of 3T3-L1 preadipocytes, the level of each 14-3-3 protein paralog is regulated independently. For instance 14-3-3β, γ, and η protein levels are increased compared to untreated cells. In contrast, 14-3-3ε protein levels decreased after differentiation while others remained constant. In silico analysis of the promoter region of each gene showed differences that explain the results obtained at mRNA and protein levels. 相似文献
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PRAS40是近几年新发现的Akt作用底物,14-3-3结合蛋白。为确定PRAS40与14-3-3蛋白7种亚基间相互作用关系,利用gateway方法构建用于酵母双杂交系统的诱饵质粒pEG-PRAS40及转录激活质粒pJG-PRAS40,将PRAS40和14-3-3各亚型质粒分别作为诱饵蛋白质粒及转录激活质粒共转化酵母细胞EGY48,通过氨基酸营养缺陷生长实验及β-半乳糖苷酶显色反应分析两种蛋白相互作用程度。酶切鉴定证实成功地构建了pEG-PRAS40和pJG-PRAS40质粒,酵母双杂交实验结果显示PRAS40可以和14-3-3亚型tau,beta,zeta及epsilon相结合,epsilon较强,beta和zeta次之,tau较弱。此结果将为深入研究PRAS40与14-3-3蛋白生物学功能及发现药物靶标奠定基础。 相似文献
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Birkenfeld J Kartmann B Anliker B Ono K Schlötcke B Betz H Roth D 《Biochemical and biophysical research communications》2003,302(3):526-533
14-3-3 proteins are ubiquitously expressed proteins which serve as central adaptors in different signal transduction cascades. In this study, yeast two-hybrid screening of a rat brain cDNA library identified a novel gene product termed zetin 1/rBSPRY that interacts with 14-3-3 zeta. The zetin 1/rBSPRY gene is ubiquitously expressed in a variety of rat tissues, with highest expression being found in testis. In adult brain, high levels of zetin 1/rBSPRY mRNA were observed in the hippocampus, cerebral cortex, and piriform cortex. Biochemical studies confirmed zetin 1/rBSPRY to interact with 14-3-3 zeta. Transient co-transfection in COS 7 cells caused a partial redistribution of zetin 1/rBSPRY into 14-3-3 zeta enriched submembranous foci at leading edges. Our results suggest a role for zetin 1/rBSPRY-14-3-3 interactions at specialized submembrane domains. 相似文献
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Summary. The 14-3-3 proteins are a family of abundant, widely expressed acidic polypeptides. The seven isoforms interact with over
70 different proteins. 14-3-3 isoforms have been demonstrated to be involved in the control of positive as well as negative
regulators of mammalian cell proliferation. Here we used the approach of inactivating 14-3-3 protein functions via overexpression
of dominant negative mutants to analyse the role of 14-3-3 proteins in mammalian cell proliferation. We found 14-3-3 dominant
negative mutants to downregulate the proliferation rates of HeLa cells. Overexpression of these dominant negative mutants
triggers upregulation of the protein levels of the cyclin-dependent kinase inhibitor p27, a major negative cell cycle regulator.
In addition, they downregulate the protein levels of the important cell cycle promoter cyclin D1. These data provide new insights
into mammalian cell proliferation control and allow a better understanding of the functions of 14-3-3 proteins. 相似文献
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hPFTAIRE1 (PFTK1), a Cdc2-related protein kinase, is highly expressed in human brain. It exhibits cytoplasmic distribution in Hela cells, although it contains two nuclear localization signals (NLSs) in its N-terminus. To search for its substrates and regulatory components, we screened a two-hybrid library by using the full-length hPFTAIRE1 as a bait. Four 14-3-3 isoforms (β,ε,η,τ) were identified interacting with the hPFTAIRE1. We found a putative 14-3-3 binding consensus motif(RHSSPSS) in the hPFTAIRE 1, which overlapped with its second NLS. Deletion of the RHSSPSS motif or substitution of Ser^119 gwithAla in the conserved binding motif abolished the specific interaction between the hPFTAIRE 1 and the 14-3 -3 proteins. The mutant S 120A hPFTAIRE1 also showed a weak interaction to the 14-3-3 proteins. The results suggested that the Ser^119 is crucial for the interaction between hPFTAIREI and the 14-3-3 proteins. All the hPFTAIRE1 mutants distributed in cytoplasm of Hela cells and human neuroblastoma cells (SH-SY5Y) when fused to the C-terminus of a green fluorescent protein (GFP), indicating that binding with the 14-3-3 proteins does not contribute to the subcellular localization of the hPFTAIRE1, although the binding may be involved in its signaling regulation. 相似文献
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