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Nucleotide sequence of the herpes simplex virus type 2 thymidine kinase gene. 总被引:21,自引:5,他引:16
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We have determined the complete nucleotide sequence of the thymidine kinase gene of herpes simplex virus (HSV) type 2 strain 333. The sequence of the thymidine kinase gene exhibits an open translational reading frame of 1,128 nucleotides encoding a protein of 376 amino acids. The DNA sequence was compared with that of the HSV type 1 thymidine kinase gene from strain MP (S. L. McKnight, Nucleic Acids Res. 8:5949-5964, 1980) and from strain CL 101 (M. J. Wagner, J. A. Sharp, and W. C. Summers, Proc. Natl. Acad. Sci. U.S.A. 78:1441-1445, 1981) to assess the extent of intra- and intertypic variation for one viral gene. The nucleotides encoding the structural gene varied 1.7% between the two HSV type 1 strains and 19% between HSV type 1 and HSV type 2, which translated to differences in the amino acid sequence of the two proteins of 1.9 and 27%, respectively. The DNA encoding the 5' regulatory sequences appeared to be more conserved than the DNA coding for the structural gene, and the DNA at the 3' end of the gene was the least homologous. 相似文献
3.
Induction of uracil-DNA glycosylase and dUTP nucleotidohydrolase activity in herpes simplex virus-infected human cells 总被引:9,自引:0,他引:9
HeLa BU cells infected with either the type 1 or the type 2 forms of herpes simplex virus show an increase in the activities of uracil-DNA glycosylase and dUTP nucleotidohydrolase. Under optimal conditions, uracil-DNA glycosylase activity increases approximately 40-fold in HSV type 2-infected cells. In herpes simplex virus (HSV) type 1-infected cells, uracil-DNA glycosylase activity increases only 6-fold. At a KCl concentration of 100 mM, uracil-DNA glycosylase derived from HSV type 2-infected cells is activated 2-fold, while the glycosylase extracted from mock infected HeLa BU cells is inhibited almost 90% at 100 mM KCl. dUTP nucleotidohydrolase activity increases 4-fold and 3-fold, respectively, in HSV type 1- and HSV type 2-infected HeLa BU cells. Nondenaturing polyacrylamide gel electrophoresis of extracts derived from the type 1- and type 2-infected cells indicates distinct electrophoretic mobilities from the host cell enzyme. dUTP nucleotidohydrolase RF values for the mock infected cells, HSV type 1, and HSV type 2 are 0.5, 0.25, and 0.33, respectively. Serum from rabbits immunized against cells infected with herpes simplex virus type 1 or type 2 specifically neutralizes the dUTPase and uracil-DNA glycosylase activities extracted from herpes simplex virus-infected cells. This serum does not neutralize dUTPase or uracil-DNA glycosylase activity derived from mock infected cells. 相似文献
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We have identified a new human LIM domain gene by isolating an autoantigenic cDNA clone from a human breast tumor cDNA library. The predicted amino acid sequence of the cDNA clone's 495 bp open reading frame contains two tandem LIM domain motifs, and within the LIM domain region there is 62% identity with the analogous region of the LIM-only gene LMO1. The homology to LMO1 is restricted to the 360 bp region encoding the tandemly repeated LIM domains, the rest of the open reading frame as well as the extensive, GC-rich 5' untranslated region, and 3' region of the 2 kb cDNA sequence are unrelated to any known genes. This gene has been designated LMO4. 相似文献
6.
Spliced RNA from the IR1-U2 region of Epstein-Barr virus: presence of an open reading frame for a repetitive polypeptide. 总被引:34,自引:6,他引:28
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We have constructed a cDNA library from the cytoplasmic RNAs of Raji cells, a Burkitt's lymphoma cell line latently infected with Epstein-Barr virus. We report here the characterization of a cDNA representing a spliced RNA transcribed from the IR1-U2 region of the viral genome. The cDNA is 1007 bp long. The 5' region contains three tandem repeats of two exons, 66 and 132 bp, which are transcribed from the IR1 repeats. The 3' region is formed from four exons transcribed from U2. An open reading frame extends from the 5' end to position 784, and includes the repeats. This reading frame presumably corresponds to the carboxy-terminal 261 amino acids of a polypeptide containing several repeats of a 66 amino acid sequence. Since it would be encoded by the IR1-U2 region of the viral genome, the putative polypeptide might be involved in the process of growth-transformation of B-lymphocytes. 相似文献
7.
B C Ossendorp G P van Heusden K W Wirtz 《Biochemical and biophysical research communications》1990,168(2):631-636
The affinity-purified antibody against rat liver non-specific lipid transfer protein (nsL-TP; sterol carrier protein 2) was used to screen a lambda-gt11 rat liver cDNA library. Positive cDNA clones were further identified by Southern blot analysis and sequenced. The largest cDNA clone consisted of 1851 bp starting at the 5' end with an open reading frame of 1545 bp. The 369 bp located at the 3' end of this open reading frame corresponded with the amino acid sequence of nsL-TP. 相似文献
8.
Lanes O Leiros I Smalås AO Willassen NP 《Extremophiles : life under extreme conditions》2002,6(1):73-86
Two distinct forms of the highly conserved uracil-DNA glycosylase (UNG) have been isolated from Atlantic cod (Gadus morhua) liver cDNA by rapid amplification of cDNA ends (RACE). From the cDNA sequences, both forms were deduced to encode an open reading frame of 301 amino acids, with an identical 267-amino-acid C-terminal region and different N-terminal regions of 34 amino acids. By comparison with the human UNG sequences, the two forms were identified as possible mitochondrial (cUNG1) and nuclear (cUNG2) forms. Several constructs of recombinant cUNG (rcUNG) were expressed in Escherichia coli in order to optimize the yield. The recombinant enzyme was purified to apparent homogeneity as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Activity and stability experiments showed that rcUNG was similar to cUNG previously purified from Atlantic cod liver, and was more pH- and temperature labile than a recombinant human UNG (rhUNG). Under optimal assay conditions for both rcUNG and rhUNG, the turnover number (k(cat)) was three times higher for rcUNG compared with rhUNG, with an identical K(M), resulting in a threefold higher catalytic efficiency (k(cat)/K(M)) for rcUNG. These activity and stability experiments reveal cold-adapted features in rcUNG. Homology models of the catalytic domains of Atlantic cod (cUNG) and mouse uracil-DNA glycosylase (mUNG) were built using the human UNG (hUNG) crystal structure as a template. The unique amino acid substitutions observed in cod UNG were mainly located in the N- and C-terminal parts of the sequence. The analysis indicated a more stable N-terminal, a more flexible C-terminal, and a less stabilized core in cUNG as compared with the mammalian UNGs. Substitution of several amino acids in or near the DNA-binding site in cUNG could give rise to a more positively charged surface and a higher electrostatic potential near the active site compared with the mammalian UNGs. The higher potential may increase the electrostatic interactions between the enzyme and DNA, and may explain the increased substrate affinity and, in combination with the higher flexibility, the higher catalytic efficiency observed for rcUNG. 相似文献
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Isolation, characterization, and expression of cDNAs encoding murine alpha-mannosidase II, a Golgi enzyme that controls conversion of high mannose to complex N-glycans 总被引:18,自引:6,他引:12
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Golgi alpha-mannosidase II (GlcNAc transferase I-dependent alpha 1,3[alpha 1,6] mannosidase, EC 3.2.1.114) catalyzes the final hydrolytic step in the N-glycan maturation pathway acting as the committed step in the conversion of high mannose to complex type structures. We have isolated overlapping clones from a murine cDNA library encoding the full length alpha-mannosidase II open reading frame and most of the 5' and 3' untranslated region. The coding sequence predicts a type II transmembrane protein with a short cytoplasmic tail (five amino acids), a single transmembrane domain (21 amino acids), and a large COOH-terminal catalytic domain (1,124 amino acids). This domain organization which is shared with the Golgi glycosyl-transferases suggests that the common structural motifs may have a functional role in Golgi enzyme function or localization. Three sets of polyadenylated clones were isolated extending 3' beyond the open reading frame by as much as 2,543 bp. Northern blots suggest that these polyadenylated clones totaling 6.1 kb in length correspond to minor message species smaller than the full length message. The largest and predominant message on Northern blots (7.5 kb) presumably extends another approximately 1.4-kb downstream beyond the longest of the isolated clones. Transient expression of the alpha-mannosidase II cDNA in COS cells resulted in 8-12-fold overexpression of enzyme activity, and the appearance of cross-reactive material in a perinuclear membrane array consistent with a Golgi localization. A region within the catalytic domain of the alpha-mannosidase II open reading frame bears a strong similarity to a corresponding sequence in the rat liver endoplasmic reticulum alpha-mannosidase and the vacuolar alpha-mannosidase of Saccharomyces cerevisiae. Partial human alpha-mannosidase II cDNA clones were also isolated and the gene was localized to human chromosome 5. 相似文献
11.
《Gene》1997,189(2):175-181
Uracil-DNA glycosylase (UDG) is the enzyme responsible for the first step in the base-excision repair pathway that specifically removes uracil from DNA. Here we report the isolation of the cDNA and genomic clones for the mouse uracil-DNA glycosylase gene (ung) homologous to the major placental uracil-DNA glycosylase gene (UNG) of humans. The complete characterization of the genomic organization of the mouse uracil-DNA glycosylase gene shows that the entire mRNA coding region for the 1.83-kb cDNA of the mouse ung gene is contained in an 8.2-kb SstI genomic fragment which includes six exons and five introns. The cDNA encodes a predicted uracil-DNA glycosylase (UDG) protein of 295 amino acids (33 kDa) that is highly similar to a group of UDGs that have been isolated from a wide variety of organisms. The mouse ung gene has been mapped to mouse chromosome 5 using fluorescence in situ hybridization (FISH). 相似文献
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Molecular cloning of human uracil-DNA glycosylase, a highly conserved DNA repair enzyme. 总被引:24,自引:3,他引:21
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Uracil-DNA glycosylase is the DNA repair enzyme responsible for the removal of uracil from DNA, and it is present in all organisms investigated. Here we report on the cloning and sequencing of a cDNA encoding the human uracil-DNA glycosylase. The sequences of uracil-DNA glycosylases from yeast, Escherichia coli, herpes simplex virus type 1 and 2, and homologous genes from varicella-zoster and Epstein-Barr viruses are known. It is shown in this report that the predicted amino acid sequence of the human uracil-DNA glycosylase shows a striking similarity to the other uracil-DNA glycosylases, ranging from 40.3 to 55.7% identical residues. The proteins of human and bacterial origin were unexpectedly found to be most closely related, 73.3% similarity when conservative amino acid substitutions were included. The similarity between the different uracil-DNA glycosylase genes is confined to several discrete boxes. These findings strongly indicate that uracil-DNA glycosylases from phylogenetically distant species are highly conserved. 相似文献
13.
Exon-intron boundary sliding in the generation of two mRNAs coding for porcine urokinase-like plasminogen activator 总被引:2,自引:0,他引:2
Y Nagamine D Pearson M Grattan 《Biochemical and biophysical research communications》1985,132(2):563-569
We have analyzed four full-length cDNA clones to porcine urokinase-like plasminogen activator (uPA) mRNA. DNA sequencing revealed a deletion of 27 nucleotides in one cDNA. The comparison of cDNA and genomic sequences indicated that this length polymorphism was due to an alternative splicing of two potential 5' splice sites to a unique 3' splice site. As the difference was 27 nucleotides (corresponding to 9 amino acids) and there was no termination codon within the same reading frame in this region, the two different mRNAs might be equally biologically active. 相似文献
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The nucleotide sequence of a full length cDNA clone encoding rat liver urate oxidase 总被引:1,自引:0,他引:1
K Alvares M R Nemali P G Reddy X D Wang M S Rao J K Reddy 《Biochemical and biophysical research communications》1989,158(3):991-995
Recently we reported the sequence of a cDNA clone (pUOX-1), isolated from a lambda gt11 cDNA library, which encoded for rat liver urate oxidase (EC 1.7.3.3), but this clone lacked the nucleotide sequences encoding the N-terminal region for this enzyme. Using the cDNA insert from the pUOX-1 clone as a probe, we have now isolated a full length cDNA clone, pUOX-2, from a lambda gt10 library by plaque hybridization. Nucleotide sequence analysis of the pUOX-2 clone showed that it has 1379 base pairs with an open reading frame coding for 303 amino acid residues corresponding to a molecular mass of 34,931 daltons. In addition to the open reading frame the pUOX-2 contains 439 bp of 3'-untranslated and 41 bp of 5'-untranslated sequences. The consensus polyadenylation signal AATAAA precedes a stretch of poly(A)+ residues at the 3' end. 相似文献
15.
Nucleotide sequence of a full-length cDNA for human fibroblast poly(ADP-ribose) polymerase 总被引:15,自引:0,他引:15
K Uchida T Morita T Sato T Ogura R Yamashita S Noguchi H Suzuki H Nyunoya M Miwa T Sugimura 《Biochemical and biophysical research communications》1987,148(2):617-622
The complete nucleotide sequence of human fibroblast poly(ADP-ribose) polymerase cDNA was determined. The cDNA contains an open reading frame for a 1014 amino acid polypeptide. In the DNA binding domain of poly(ADP-ribose) polymerase, there are predicted alpha-helix-turn-alpha-helix structures and two sequences each of about 100 amino acids that are similar to each other containing potential cysteine-zinc DNA binding structures. Within the 3' untranslated region, there is an AT-rich sequence containing ATTTA, a possible mRNA destabilizer. 相似文献
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Sequence of 3,687 nucleotides from the 3' end of Sendai virus genome RNA and the predicted amino acid sequences of viral NP, P and C proteins. 总被引:30,自引:9,他引:30
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T Shioda Y Hidaka T Kanda H Shibuta A Nomoto K Iwasaki 《Nucleic acids research》1983,11(21):7317-7330
The sequence of 3,687 nucleotides from the 3' end of the Sendai virus genome (Z strain) was determined by a molecular cloning technique followed by rapid sequence analysis. Two large open reading frames, one consisting of 1,572 nucleotides and the other of 1,704 nucleotides, were observed in the region, that is OP-1 and OP-2 from the 3' end of the genome. The amino acid sequences of the gene products were predicted from the observed sequence. Determination of amino acid compositions of viral proteins, P, HN, Fo, NP and M, led us to conclude that NP and P are the gene products of OP-1 and OP-2, respectively. An additional open reading frame consisting of 612 nucleotides (OP-3) was discovered in the 3' most proximal region of OP-2. The predicted product of OP-3 was considered to be viral non-structural protein C. The leader sequence of 51 nucleotides at the 3' terminal of the genome and consensus sequences at 3' and 5' ends of each gene for proteins NP and P were identified. 相似文献
18.
Isolation and characterization of bovine and mouse terminal deoxynucleotidyltransferase cDNAs expressible in mammalian cells. 总被引:15,自引:5,他引:10
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O Koiwai T Yokota T Kageyama T Hirose S Yoshida K Arai 《Nucleic acids research》1986,14(14):5777-5792
We have isolated nearly full-length cDNA clones of terminal deoxynucleotidyltransferase (TdT) from calf thymus and mouse lymphoma cDNA libraries. The libraries were constructed using the pcD vector system which permits the expression of cDNA inserts in mammalian cells. The bovine TdT cDNA clone contains an open reading frame coding for 520 amino acids, Mr 59,678. The mouse TdT cDNA clone contains an open reading frame of 1,587 bp, whose translated cDNA encodes a 60,004 dalton protein. The mouse TdT cDNA clone contains 60 bp in the 3' end region of the coding sequence not found in the bovine TdT cDNA sequence, otherwise, the clones share about 80% homology. A possible nuclear-localization-sequence (Pro-Arg-Lys-Lys-Arg-Pro-Arg) was conserved in the N-terminal region in the mouse and bovine cDNA clones. Bovine and mouse cDNAs transfected into COS7 monkey fibroblasts directed the synthesis of enzymatically active protein of Mr 60,000 which was detected immunologically using polyclonal rabbit antibody against bovine TdT. Bovine TdT expressed in COS7 cells by nearly full-length cDNA clone was localized in the nucleus and the translational product of pOK103 lacking the nuclear-localization-sequence was localized in the cytoplasm. 相似文献
19.
Federico Focher Annalisa Verri Simona Verzeletti Paolo Mazzarello Silvio Spadari 《Chromosoma》1992,102(1):S67-S71
We have recently demonstrated that mammalian uracil-DNA glycosylase activity is undetectable in adult neurons. On the basis of this finding we hypothesized that uracil, derived either from oxidative deamination of cytosine or misincorporation of dUMP in place of dTMP during DNA repair by the unique nuclear DNA polymerase present in adult neurons, DNA polymerase β, might accumulate in neuronal DNA. Uracil residues could also arise in the herpes simplex 1 (HSV1) genome during latency in nerve cells. We therefore suggest a role for the virus encoded uracil-DNA glycosylase in HSV1 reactivation and in the first steps of DNA replication. We show here 1) that the viral DNA polymerase incorporates dUTP in place of dTTP with a comparable efficiencyin vitro; 2) that virus specific DNA/protein interactions between the virus encoded origin binding protein and its target DNA sequence is altered by the presence of uracil residues in its central region TCGCA. Thus uracil, present in viral OriS or other key sequences could hamper the process leading to viral reactivation. Hence, HSV1 uracil-DNA glycosylase, dispensable in viral proliferation in tissue culture, could be essential in neurons for the “cleansing” of the viral genome of uracil residues before the start of replication. 相似文献