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Pro-apoptotic ASY/Nogo-B protein associates with ASYIP 总被引:3,自引:0,他引:3
Qi B Qi Y Watari A Yoshioka N Inoue H Minemoto Y Yamashita K Sasagawa T Yutsudo M 《Journal of cellular physiology》2003,196(2):312-318
We have previously shown that ectopic expression of the ASY/Nogo-B gene induced apoptosis in various cancer cell lines. Nogo-A, a splice variant of the ASY, has been reported to have an inhibitory effect on neuronal regeneration in the central nervous system. To investigate the mechanism of ASY-induced apoptosis or inhibition of neuronal regeneration, we cloned a cDNA for the ASY-interacting protein from the human cDNA library using the yeast two-hybrid method, and obtained a cDNA we designated as ASYIP. The ASYIP protein contains two hydrophobic regions and a double lysine endoplasmic reticulum (ER) retrieval motif at its C-terminus, which was shown to be identical to RTN3, a reticulon family protein of unknown function. We showed that ASY and ASYIP proteins formed a complex also in human cells. Mutational analysis indicated that both of the hydrophobic regions of the ASYIP protein were required for the association. By immunofluorescence analysis, the ASYIP protein was shown to be co-localized with ASY in the ER. Characterization of the ASYIP gene may be very useful in clarifying the mechanism of ASY-induced apoptosis or Nogo-involved inhibition of neuronal regeneration in the central nervous system. 相似文献
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Tetsunori Seki 《Immunogenetics》1989,30(1):5-12
Two varieties of similar, but structurally distinct, cDNA clones for the human low-affinity receptors for the Fc portion of
immunoglobulin G (FcγRII) have been isolated. One type of clone was obtained from human B lymphocytes, and the other from
PHA-activated peripheral T cells and monocytes. Transfection of both prototype clones into Cos-7 cells and subsequent specific
staining with monoclonal antibodies of the CDw32 group confirmed the identification of the gene products. The nucleotide sequence
of the cDNA clone from B lymphocytes contains an open reading frame that encodes a protein of relative mass (M
r) 27000 with an extracellular domain of 179 amino acids containing three potential N-glycosylation sites, a 26 amino acid
transmembrane domain, and a 44 amino acid cytoplasmic domain. The clones from peripheral T cells and monocytes both encoded
a protein ofM
r 31000 with a 179 amino acid extracellular domain containing two potential N-glycosylation sites and a 26 amino acid transmembrane
domain. The two types of clones had similar sequences in their immunoglobulin-like extracellular and transmembrane domains,
but differed in their leader sequences and 3′-untranslated regions. The most notable difference between the clones was the
presence of a distinctive 76 amino acid cytoplasmic domain in those isolated from T cells and monocytes. 相似文献
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Cloning and characterization of the 14-3-3 protein gene from the halotolerant alga Dunaliella salina
Previous studies have demonstrated that 14-3-3 proteins exist in all the eukaryotic organisms studied; however, studies on
the 14-3-3 proteins have not been involved in the halotolerant, unicellular green alga Dunaliella salina so far. In the present study, a cDNA encoding 14-3-3 protein of D. salina was cloned and sequenced by PCR and rapid amplification of cDNA end (RACE) technique based on homologous sequences of the
14-3-3 proteins found in other organisms. The cloned cDNA of 1485 bp in length had a 29.2 kDa of molecular weight and contained
a 774 bp of open reading frame encoding a polypeptide of 258 amino acids. Like the other 14-3-3 proteins, the deduced amino
acid sequences of the D. salina 14-3-3 protein also contained two putative phosphorylation sites within the N-terminal region (positions 62 and 67). Furthermore,
an EF hand motif characteristic for Ca2+-binding sites was located within the C-terminal part of this polypeptide (positions 208–219). Analysis of bioinformatics
revealed that the 14-3-3 protein of D. salina shared homology with that of other organisms. Real-time quantitative PCR demonstrated that expression of the 14-3-3 protein
gene is cell cycle-dependent. 相似文献
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The gene encoding a cuticle-degrading serine protease was cloned from three isolates of Lecanicillium psalliotae (syn. Verticillium psalliotae) by 3′ and 5′ RACE (rapid amplification of cDNA ends) method. The gene encodes for 382 amino acids and the protein shares
conserved motifs with subtilisin N and peptidase S8. Comparison of translated cDNA sequences of three isolates revealed one
amino acid polymorphism at position 230. The deduced protease sequence shared high degree of similarities to other cuticle-degrading
proteases from other nematophagous fungi. 相似文献
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果蝇程序化死亡基因5(PDCD5)同源cDNA的克隆和序列分析 总被引:2,自引:0,他引:2
为了解人类白血病细胞凋亡相关新基因 TFAR1 9(PDCD5,programmed cell death5)在不同种属间的序列同源性 ,利用 EST(expression sequence tag)拼接、RT- PCR、DNA序列测定技术及计算机分析技术 ,首次成功地进行了果蝇 PDCD5同源 c DNA编码区基因克隆和序列分析 .发现果蝇与小鼠及果蝇与人 PDCD5在核苷酸水平上分别有 57.5%和 57.1 %的同源性 ,在氨基酸水平上分别有 46.8%和 46.4%的同源性 .功能区分析发现 ,果蝇 PDCD5c DNA编码 1 33个氨基酸 ,计算机预测可能是一种核蛋白 ,含 5个可能的酪蛋白激酶 (casein kinase )磷酸化位点 ,2个可能的 PKC磷酸化位点 ,与人 PDCD5的功能区类似 .因而果蝇 PDCD5是与人 PDCD5同源的新基因 ,可能都与细胞程序化死亡相关 . 相似文献
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Cloning and expression of a gene for an alpha-glucosidase from Saccharomycopsis fibuligera homologous to family GH31 of yeast glucoamylases 总被引:4,自引:0,他引:4
Cloning of cDNA encoding an α-glucosidase from the dimorphous yeast Saccharomycopsis fibuligera and characterization of the gene product were performed. The cDNA of the putative α-glucosidase gene consists of 2,886 bp, which includes an open reading frame encoding a 19 amino acid signal peptide at the N-terminal end and a 944 amino acid mature protein with a predicted molecular mass of 105.4 kDa and pI value of 4.52. The deduced amino acid sequence shows a high degree of identity (70%) with two yeast glucoamylases, namely, the extracellular glucoamylase Gam from Schwanniomyces occidentalis and the cell surface glucoamylase Gca from Candida albicans. The recombinant product, synthesized in Saccharomyces cerevisiae, is localized on the cell surface and hydrolyses maltooligosaccharides exclusively without the ability to digest soluble starch, which is consistent with the specificity characteristic of α-glucosidase, EC. 3.2.1.20. 相似文献
10.
Cloning, sequencing and expression of cDNA encoding growth hormone from Indian catfish (Heteropneustes fossilis) 总被引:2,自引:0,他引:2
A tissue-specific cDNA library was constructed using polyA+ RNA from pituitary glands of the Indian catfishHeteropneustes fossilis (Bloch) and a cDNA clone encoding growth hormone (GH) was isolated. Using polymerase chain reaction (PCR) primers representing
the conserved regions of fish GH sequences the 3′ region of catfish GH cDNA (540 bp) was cloned by random amplification of
cDNA ends and the clone was used as a probe to isolate recombinant phages carrying the full-length cDNA sequence. The full-length
cDNA clone is 1132 bp in length, coding for an open reading frame (ORF) of 603 bp; the reading frame encodes a putative polypeptide
of 200 amino acids including the signal sequence of 22 amino acids. The 5′ and 3′ untranslated regions of the cDNA are 58
bp and 456 bp long, respectively. The predicted amino acid sequence ofH. fossils GH shared 98% homology with other catfishes. Mature GH protein was efficiently expressed in bacterial and zebrafish systems
using appropriate expression vectors. The successful expression of the cloned GH cDNA of catfish confirms the functional viability
of the clone. 相似文献
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Hua Xie Peirong Xu Yanlong Jia Jie Li Yumin Lu Lexun Xue 《Journal of applied phycology》2007,19(5):497-504
A cDNA corresponding to the nitrate reductase (NR) gene from Dunaliella salina was isolated by RT-PCR and (5′/3′)-RACE techniques. The full-length cDNA sequence of 3,694 bp contained an open reading frame
of 2,703 bp encoding 900 amino acids, a 5′-untranslated region of 151 bp and a 3′-untranslated sequence of 840 bp with a poly
(A) tail. The putative gene product exhibited 78%, 65%, 59% and 50% identity in amino acid sequence to the corresponding genes
of Dunaliella tertiolecta, Volvox carteri, Chlamydomonas reinhardtii, and Chlorella vulgaris, respectively. Phylogenetic analysis showed that D. salina NR clusters together with known NR proteins of the green algae. The molecular mass of the encoded protein was predicted to
be 99.5 kDa, with an isoelectric point of 8.31. This protein shares common structural features with NRs from higher plants
and green algae. The full-length cDNA was heterologously expressed in Escherichia coli as a fusion protein, and accumulated to up to 21% of total bacteria protein. Recombinant NR protein was active in an enzyme
assay, confirming that the cloned gene from D. salina is indeed NR. 相似文献
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François Barrieu Dominique Thomas Danièle Marty-Mazars Maryse Charbonnier Francis Marty 《Planta》1998,204(3):335-344
The vacuolar membrane (tonoplast) of plant cells contains aquaporins, protein channels that facilitate the selective transport
of water. These tonoplast intrinsic proteins (TIPs) of 23–29 kDa belong to the ancient major intrinsic protein (MIP) family.
A monospecific polyclonal antiserum directed against a 26 kDa intrinsic protein from the tonoplast of meristematic cells from
cauliflower (Brassica oleracea L. var. botrytis) was used to screen a cDNA library. Two distinct cDNAs have been isolated. Both clones, c26-1 and c26-2, encode closely related TIPs. The c26-1 insert, consisting of 933 bp upstream of the poly(A) tail, is a full-length cDNA with an open reading frame encoding a protein
of 251 amino acids with a calculated Mr of 25 500. The c26-2 insert is a 5′ truncated cDNA. The two cDNAs share 90.5% sequence identity within their overlapping coding regions but only
35% sequence identity in the 3′␣untranslated regions, indicating that highly related TIP-encoding genes are expressed in meristematic
cells. Although TIPs have previously been found in a variety of cell types, they have not been found in meristems. The derived
amino acid sequences (BobTIP26-1 and BobTIP26-2, respectively) closely resemble the aquaporin γ-TIP from Arabidopsis thaliana. Northern blot analysis and in situ hybridization show that BobTIP26 mRNAs preferentially accumulate in highly meristematic cells, mostly before and during cell enlargement, and in the living
cells of the xylem. This differential pattern of expression is also found by immunodetection of BobTIP26 polypeptides. The
gene expression patterns are discussed with respect to the probable function of the gene products.
Received: 27 March 1997 / Accepted: 20 May 1997 相似文献
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Liang Wei Yi Cao Linhan Bai Xue Liang Tingting Deng Jing Li Dairong Qiao 《Journal of applied phycology》2007,19(1):89-94
Genes encoding proteins of the major light-harvesting complex of photosystem II (LHCII) in higher plants are well studied.
However, little is known about the corresponding genes in the green alga Dunaliella salina, although this knowledge might provide valuable information about the respective roles of each LHCII protein at the molecular
level under extreme environmental conditions. Here, we describe an additional LhcII gene from D. salina. An LhcII cDNA cloned by screening a D. salina cDNA library contains an open reading frame encoding a protein of 261 amino acids with a calculated molecular mass of 27.8 kDa.
The deduced amino acid sequence shows high homology with other LHCII proteins. Genomic DNA—obtained by PCR using a specific
primer set corresponding to the 5′ and 3′ untranslated regions—was used to determine the intron-exon structure. Short-term
changes in mRNA levels after a shift from low-light to high-light or dark conditions were analyzed by real-time quantitative
PCR, and indicated that this gene expresses different mRNA levels under different light conditions. 相似文献
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Md. Maksudul Alam Sazia Sharmin Zinnatun Nabi Shakhinur Islam Mondal Md. Shahidul Islam Sarmah Bin Nayeem Muhammad Shoyaib Haseena Khan 《Plant Molecular Biology Reporter》2010,28(3):394-402
A putative leucine-rich repeat receptor-like protein kinase (LRR-RLK) gene together with its 5′ and 3′ untranslated regions
of jute (Corchorus olitorius L.) has been identified and sequenced. The gene is 3,371 bp long containing two exons and one intron. The coding sequence
of the gene is 2,879 bp long encoding a peptide of 957 amino acids. The predicted protein contains several domains and motifs
characteristic of a transmembrane protein kinase. It is complete with domains for an N-terminal leucine-rich repeat and a
protein kinase core, an active site for serine/threonine protein kinase, an ATP binding conserved site and a transmembrane
region. Expression of the gene is induced by low temperature, high salt concentration, dehydration, abscisic acid treatment,
and fungal infection, suggesting the involvement of the gene in multiple stress response pathways in jute (C. olitorius L.). A possible mechanism of the role of the gene in signal transduction and environmental stress response is discussed.
To date, LRR-RLK is the only jute gene which has been completely sequenced and characterized. 相似文献
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Szyszka R 《Folia microbiologica》1999,44(2):142-152
Phosphorylation of ribosomal acidic proteins ofSaccharomyces cerevisiae is an important mechanism regulating a number of active ribosomes. The key role in the regulatory mechanism is played by
specific phosphoprotein kinases and phosphoprotein phosphatases. Three different cAMP-independent protein kinases phosphorylating
acidic ribosomal proteins have been identified and characterized. The protein kinase 60S (PK60S), RAP kinase, and casein kinase
type 2 (CK2). All three protein kinases phosphorylate serine residues which are localized in the C-terminal end of phosphoproteins.
Synthetic peptides were used to determinate the amino acid sequence of phosphoacceptor site for PK60S. Peptide AAEESDDD derived
from phosphoproteins YP1β/β′ and YP2α turned out to be the best substrate for PK60S. A number of halogenated benzimidazoles
and 2-azabenzimidazoles were tested as inhibitors of the three protein kinases. 4,5,6,7-Tetrabromo-2-azabenzimidazole inhibits
phosphorylation only of these polypeptides phosphorylated by protein kinase 60S, namely YP1β/β′ and YP2α, but not the other,
YP1α and YP2β phosphorylated by protein kinases RAP and CK2. RAP kinase has been found in an active form in the soluble fraction
ofS. cerevisiae. The enzyme uses ATP as a phosphate donor and is less sensitive to heparin than casein kinase 2. RAP kinase monophosphorylates
the four acidic proteins. The ribosome-bound proteins are a better substrate for the enzyme. Multifunctional CK2 kinase phosphorylate
all four acidic proteins. The kinase phosphorylates preferentially serine or threonine residues surrounded by cluster of acidic
residues. The enzyme activity is stimulatedin vitro by the presence of polylysine and inhibited by heparin.
Presented at theSymposium on Regulation of Translation of Genetic Information by Protein Phosphorylation, 21 st Congress of the Czechoslovak
Society for Microbiology, Hradec Králové (Czech Republic), September 6–10, 1998. 相似文献
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M. E. G. Suykerbuyk P. J. Schaap H. Stam W. Musters J. Visser 《Applied microbiology and biotechnology》1995,43(5):861-870
Rhamnogalacturonase was purified from culture filtrate ofAspergillus aculeatus after growth in medium with sugar-beet pulp as carbon source. Purified protein was used to raise antibodies in mice and with
the antiserum obtained a gene coding for rhamnogalacturonase (rhgA) was isolated from a λ cDNA expression library. The clonedrhgA gene has an open-reading frame of 1320 base pairs encoding a protein of 440 amino acids with a predicted molecular mass of
45 962 Da. The protein contains a potential signal peptidase cleavage site behind Gly-18 and three potential sites forN-glycosylation. Limited homology withA. niger polygalacturonase amino acid sequences is found. A genomic clone ofrhgA was isolated from a recombinant phage λ genomic library. Comparison of the genomic and cDNA sequences revealed that the coding
region of the gene is interrupted by three introns. Furthermore, amino acid sequences of four different peptides, derived
from purifiedA. aculeatus rhamnogalacturonase, were also found in the deduced amino acid sequence ofrhgA.A. aculeatus strains overexpressing rhamnogalacturonase were obtained by cotransformation using either theA. niger pyrA gene or theA. aculeatus pyr A gene as selection marker. For expression of rhamnogalacturonase inA. awamori theA. awamori pyrA gene was used as selection marker. Degradation patterns of modified hairy regions, determined by HPLC, show the recombinant
rhamnogalacturonase to be active, and the enzyme was found to have a positive effect in the apple hot-mash liquefaction process. 相似文献
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Rieske protein gene in the Pacific oyster Crassostrea gigas was obtained by in silico cloning for the first time, and its expression profiles and subcellular localization were determined,
respectively. The full-length cDNA of Cgisp is 985 bp in length and contains a 5′- and 3′-untranslated regions of 35 and 161 bp, respectively, with an open reading frame
of 786 bp encoding a protein of 262 amino acids. The predicted molecular weight of 30 kDa of Cgisp protein was verified by prokaryotic expression. Conserved Rieske [2Fe–2S] cluster binding sites and highly matched-pair tertiary
structure with 3CWB_E (Gallus gallus) were revealed by homologous analysis and molecular modeling. Eleven putative SNP sites and two conserved hexapeptide sequences,
box I (THLGC) and II (PCHGS), were detected by multiple alignments. Real-time PCR analysis showed that Cgisp is expressed in a wide range of tissues, with adductor muscle exhibiting the top expression level, suggesting its biological
function of energy transduction. The GFP tagging Cgisp indicated a mitochondrial localization, further confirming its physiological
function. 相似文献