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31.
Assignment of the 13C nuclear magnetic resonance spectrum of a short DNA-duplex with 1H-detected two-dimensional heteronuclear correlation spectroscopy. 总被引:1,自引:1,他引:0
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Proton-detected 1H-13C heteronuclear correlated spectroscopy [( 1H,13C]-COSY) was used to establish relations between the carbon-13 and proton nuclear magnetic resonance chemical shifts in the hexadeoxynucleoside pentaphosphate d-(GCATGC)2. Using the previously established sequence-specific proton NMR assignments, sequence-specific assignments were thus obtained for nearly all proton-bearing carbons. This approach offers a new criterion for distinguishing between the proton NMR lines of purines and pyrimidines, based on the different proton-carbon-13 coupling constants. Furthermore, the adenine ring carbon 2 has a unique carbon-13 chemical shift, which enables a straightforward identification of the adenine C2H resonances by [1H,13C]-COSY. 相似文献
32.
Application of an immunoprecipitation procedure to the study of SV40 tumor antigen interaction with mouse genomic DNA sequences. 总被引:3,自引:1,他引:2
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Simian Virus 40 (SV40) large T antigen is a DNA binding protein with high affinity for segments of the viral genome. To find out whether T antigen also binds to sequences of genomic cellular DNA we mixed T antigen and SAU 3 A restricted mouse DNA under stringent DNA binding conditions. Resulting protein-DNA complexes were immunoprecipitated using T antigen specific monoclonal or polyclonal antibodies. The DNA fragments in the immunoprecipitates were cloned in plasmid vectors. Four plasmid clones were selected for a detailed investigation of the inserted mouse DNA fragments. Nucleotide sequencing and DNase I footprint experiments showed that T antigen binds to sites in these fragments consisting of two tandemly oriented G(A)AGGC pentamers separated by AT rich spacers of different lengths. The cellular binding sites are very similar in their architecture to the SV40-DNA binding site I. The isolated cellular DNA fragments with T antigen binding sites occur only once or a few times in the mouse genome. Our data help to further define the structure of T antigen's DNA binding sites. The genetic functions of the isolated cellular DNA elements are not known. 相似文献
33.
34.
Endothelioma cells expressing the polyoma middle T oncogene induce hemangiomas by host cell recruitment 总被引:25,自引:0,他引:25
Mouse endothelioma cells expressing the polyoma middle T oncogene induced hemangiomas in a variety of species such as mice, rats, chicks, and quails. In embryos and newborn mice the hemangiomas expanded within 10-18 hr of injection, disrupting the vasculature and causing the death of the animal. In contrast, the hemangiomas formed a stable structure reminiscent of benign human hemangiomas in adult mice within 5 days. Analysis of the cells comprising the hemangioma revealed that over 95% of the endothelial cells were host derived. No induction of host cell proliferation was detected, and no endothelial mitogens were secreted by the endothelioma cells in vitro. The maintenance of the hemangioma appeared to require the continuous presence of endothelioma cells. The results indicate that these endothelioma cells act as a potent stimulating agent in the rapid formation of hemangiomas by recruiting nonproliferating host endothelial cells. 相似文献
35.
Summary The cavernous body of green monkeys contains many unmyelinated and few myelinated axons. The unmyelinated axons form terminals in the adventitia of the arteries, between trabecular muscle cells, in the interstitium, and close to endothelium cells of the sinuses. All terminals displayed predominantly small clear vesicles and very few large granular vesicles; small granular vesicles were not seen. However, in rabbit penises, terminals with many large granular vesicles are prominent. Immunohistochemistry (PAP technique) showed a dense network of VIP- and NPY-reactive fibres around the arteries and around trabecular muscles. The density of nerve fibres was particularly high around the subendothelial cushions of the helicine arteries. Double staining for NPY and VIP revealed that both peptides were colocalized. Immunocytochemistry (preembedding PAP technique) showed VIP- and NPY-reactivity in terminals with small clear vesicles; the reaction product was bound to the cytoplasmic face of different membrane types. Although the intracellular localization of the reaction product is probably due to artefactual displacement during preparation, the uniformity of the terminals questions the view that large and small granular vesicles in all species characterize peptidergic and noradrenergic terminals, respectively. The essential findings can be summarized as (1) a high degree of uniformity of nerve terminals, (2) colocalization of VIP and NPY, (3) heavy innervation of the subendothelial cushions of the helicine arteries, and (4) possible innervation of endothelial cells. 相似文献
36.
Isolation, characterization and immunological determination of basement membrane-associated heparan sulfate proteoglycan 总被引:2,自引:0,他引:2
B Olgem?ller E Schleicher A Nerlich E M Wagner K D Gerbitz 《Biological chemistry Hoppe-Seyler》1989,370(12):1321-1329
Basement membrane-associated heparan sulfate proteoglycan (HSPG) was extracted from isolated porcine glomerular basement membranes and purified by ion-exchange chromatography. The proteogycan was characterized by specific enzymatic digestions, by amino-acid analysis, by SDS-polyacrylamide gel electrophoresis and by density gradient centrifugation. Polyclonal antibodies were raised against the purified HSPG in rabbits. Antibodies were characterized by enzyme immunoassays, immunoprecipitation and immunohistological methods. They were shown to recognize specifically the core protein of HSPG from porcine, human and rat glomerular basement membrane but did not recognize HSPG from guinea pig or rabbit kidney. The affinity-purified antibodies did not cross-react with other basement membrane proteins like laminin, fibronectin or collagen type IV nor with chondroitin sulfate-rich or keratan sulfate-rich proteoglycans from human or bovine tissue. Using these antibodies an enzyme immunoassay was developed for determination of HSPG in the range of 1-100 ng/ml. Studies with cultured porcine endothelial cells showed that subendothelial basement membrane-associated HSPG may be determined with the enzyme immunoassay. 相似文献
37.
In a case of CML with a variant Philadelphia translocation (Ph1 or Ph) t(22;22) (q11;q13) in bone marrow cells and unstimulated peripheral blood cells, no cytogenetically detectable involvement of chromosome 9 was observed. Southern blot experiments using probes specific for bcr and c-sis however revealed rearrangement of the bcr, but not of PDGFB (c-sis) gene. Northern blot analysis of bone marrow RNA showed a very weak signal with the c-sis probe, while in a lymph-node biopsy PDGFB m-RNA could not be detected. Chromosomal in situ hybridization gave evidence for translocation of c-abl from chromosome 9 to Ph and of PDGFB from chromosome 22 to chromosome 9, as the result of a threefold translocation t(9;22;22). 相似文献
38.
Calcium regulates the rate of rhodopsin disactivation and the primary amplification step in visual transduction 总被引:3,自引:0,他引:3
The kinetics of the light-induced activation of transducin as well as the subsequent disactivation process can be monitored by means of a specific light scattering transient PA. In this communication it is demonstrated that the rate of transducin disactivation is calcium dependent, increasing when the calcium concentration is decreased. As a consequence of the accelerated recovery in low calcium, the time to the peak of the transducin activation process is shortened and the gain of the primary amplification step, i.e. the number of transducin molecules activated per bleached rhodopsin, is reduced. Experiments using hydroxylamine as an artificial quencher of rhodopsin activity suggest that calcium acts upon rhodopsin kinase and not upon the rate of the GTPase. This would indicate that calcium may control visual adaptation not only by regulating guanine cyclase activity, but also by affecting the primary step in the transduction cascade, the rhodopsin-transducin coupling. 相似文献
39.
A procedure for the determination of inositol by reversed-phase HPLC is described which is based on a precolumn benzoylation and detection at 230 nm. This procedure was used to assay the activity of L-myo-inositol 1-phosphate synthetase (EC 5.5.1.4) after treatment of the enzymatic product by a phosphatase. 相似文献
40.
K Hubrich-Kühner H J Buhk H Wagner H Kr?ger D Simon 《Biochemical and biophysical research communications》1989,160(3):1175-1182
The eukaryotic DNA cytosine-5-methyltransferase (E.C.2.1.1.37) is known to methylate cytosine in DNA mainly, but not exclusively in C-G. In the present study the minor, non-C-G recognition sequences of a rat DNA methyltransferase were analyzed by Maxam-Gilbert sequencing of in vitro methylated SV40 DNA. The enzyme methylates C-A and C-T at a 50-fold lower initial rate than C-G. Methylation of C-C at the 5'C was not observed in the piece of DNA sequenced. The methylation of C-A is very low in the trinucleotides ACA and CAC, the other C-A containing trinucleotides in DNA are much better methylacceptors. C-T was found methylated predominantly in the sequences CCTAA, ACTAA, and ACTGT. A comparison of the activity with different substrates is in favour of the enzyme making its recognition in the major groove of the DNA. 相似文献