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11.
Autonomous DNA replication in human cells is affected by the size and the source of the DNA. 总被引:31,自引:13,他引:18 下载免费PDF全文
We previously developed short-term and long-term assays for autonomous replication of DNA in human cells. This study addresses the requirements for replication in these assays. Sixty-two random human genomic fragments ranging in size from 1 to 21 kb were cloned in a prokaryotic vector and tested for their replication ability in the short-term assay. We found a positive correlation between replication strength and fragment length, indicating that large size is favored for efficient autonomous replication in human cells. All large fragments replicated efficiently, suggesting that signals which can direct the initiation of DNA replication in human cells are either very abundant or have a low degree of sequence specificity. Similar results were obtained in the long-term assay. We also used the same assays to test in human cells a random series of fragments derived from Escherichia coli chromosomal DNA. The bacterial fragments supported replication less efficiently than the human fragments in the short-term and long-term assays. This result suggests that while the sequence signals involved in replication in human cells are found frequently in human DNA, they are uncommon in bacterial DNA. 相似文献
12.
Behaviors of southern red oak hemicelluloses and lignin in a mild sulfuric acid hydrolysis 总被引:1,自引:0,他引:1
Removal and modification of southern red oak hemicelluloses and lignin in a 0.05%(w/v) sulfuric acid hydrolysis were investigated. The hydrolysis profile was to raise the reaction from room temperature to 150 degrees C for in 38 min and to extend the hydrolysis at 150 degrees C for 1 h. At the end of the hydrolysis, 25.5% of red oak components were dissolved, of which 58% was xylose and 17% lignin. As the hydrolysis proceeded from room temperature to 150 degrees C, a part of red oak xylan was removed to yield an oligomer fraction having maximal yield and average molecular weight of 3460 at 150 degrees C. This fraction and the bulk xylan extracted during the first 30 min at 150 degrees C were further degraded to give a lower molecular weight oligomer fraction, of which the yield and average molecular weight (2610) were highest at the end of the bulk removal of xylan. Red oak lignin, syringyl and guaiacyl units in particular, was increasingly removed with the progress of the hydrolysis. Lignin derivatives and a part of red oak extractives soluble in the hydrolysate were identified. 相似文献
13.
Antithrombin III Basel. Identification of a Pro-Leu substitution in a hereditary abnormal antithrombin with impaired heparin cofactor activity 总被引:6,自引:0,他引:6
Antithrombin III Basel is a hereditary abnormal antithrombin with normal progressive inhibition activity (normal reactive site) and reduced heparin cofactor activity (impaired heparin binding site). Structures of antithrombin III Basel and normal antithrombin III isolated from the same patient were compared by peptide mapping using the dimethylaminoazobenzene isothiocyanate precolumn derivatization technique. Of the approximately 50 tryptic peptides of normal and abnormal antithrombin III, one peptide comprising residues 40-46 had a different retention time in reversed-phase high performance liquid chromatography. The amino acid sequence of the peptide from antithrombin III Basel had a single substitution of Pro (normal) by Leu (abnormal) at position 41. This substitution is close to an Arg (residue 47) and a Trp (residue 49) which have previously been shown to be critical for heparin binding by antithrombin III. Although additional amino acid substitutions in antithrombin III Basel cannot be ruled out, this Pro-Leu replacement could cause a conformational change by increasing both the helical structure and the hydrophobicity around residue 41. These data suggest that: (i) the heparin binding site of antithrombin III encompasses the region containing residues 41, 47, and 49; and (ii) the impaired heparin cofactor activity of antithrombin III Basel is likely due to a conformational change of the heparin binding site induced by the Pro-Leu substitution at position 41. 相似文献
14.
The fluorescence lifetime and rotational correlation time of the tryptophan residue in melittin, as both a monomer and tetramer, have been measured between pH 6 and 11. The fluorescence decays are non-exponential and give lifetimes of 0.7±0.1 ns and 3.1±0.1 ns. This emission is consistent with a model in which the tryptophan residue is in slightly different environments in the protein. In a dilute solution of monomer the mean fluorescence lifetime is 2.3±0.1 ns, below pH 10, but falls to 1.7 ns at higher pH. In contrast, the melittin tetramer has a mean fluorescence lifetime of only 2.2 ns at pH 6, which falls to 1.9 ns by pH 8, and falls again above pH 10 to the same value as in monomeric melittin. The behaviour between pH 6 and 8 is explained as the quenching of the Trp residue by lysine groups, which are near to the Trp in the tetramer but in the monomer, are too distant to quench. Fluorescence anisotropy decays show that the Trp residue has considerable freedom of motion and the range of wobbling motion is 35±10° in the tetramer 相似文献
15.
This study describes the synthesis of prostaglandins (PGs) by the vascular structures of the inner ear (lateral wall = stria vascularis and spiral ligament) in vitro. The main PGs produced were PGI2, PGF2 alpha and PGE2. PGI2 and PGF2 alpha were also found in the perilymph. A 350 mg/kg ip injection of aspirin decreased PG synthesis by the lateral wall and PG levels in perilymph. This effect was reversed after 3 days. Gentamicin (10(-9) to 10(-5) M) decreased significantly and reversibly PG synthesis in vitro, as did 100 mg/kg ip injection. Acoustic stimulation increased ex vivo PGI2 and PGE2 synthesis without modifying PG levels in perilymph. Results suggest that PGs could be one humoral mediator of the cochlear microcirculation homeostasis, and, possibly, of the circulatory disturbances reported after acoustic stimulation. The decreased PG synthesis after gentamicin treatment could account for the angiotoxic component observed in aminoglycoside ototoxicity. 相似文献
16.
Concentrations of high-mobility-group proteins in the nucleus and cytoplasm of several rat tissues 总被引:6,自引:0,他引:6
Nuclear and cytoplasmic fractions were isolated from various tissues of the rat by a nonaqueous technique. The high-mobility-group (HMG) proteins were extracted from these fractions with acid and separated by one- and two-dimensional PAGE. The concentrations of high-mobility-group proteins HMG1, HMG2, and HMG17 in the nucleus and cytoplasm were then estimated from the staining intensities of the electrophoretic bands. The cytoplasmic concentrations of these proteins were very low--usually less than 1/30 of those present in the corresponding nuclear fractions. For the tissues studied (liver, kidney, heart, and lung), the concentrations of HMG proteins in the nucleus did not differ significantly from one tissue to another. Averaged over the four tissues investigated, there were 0.28 molecule of HMG1, 0.18 molecule of HMG2, and 0.46 molecule of HMG17 per nucleosome. These values are considerably higher than those that have been reported previously. 相似文献
17.
The mechanism of biliary secretion of reduced glutathione. Analysis of transport process in isolated rat-liver canalicular membrane vesicles 总被引:7,自引:0,他引:7
Transport of reduced glutathione (GSH) was studied in isolated rat liver canalicular membrane vesicles by a rapid filtration technique. The membrane vesicles exhibit uptake of [2-3H]glycine--labeled GSH into an osmotically reactive intravesicular space. Although the canalicular membrane vesicles possess gamma-glutamyltransferase and aminopeptidase M, enzymes that hydrolyze glutathione into component amino acids, inactivation of the vesicle-associated transferase by affinity labeling with L-(alpha S,5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (AT-125) had no effect on the initial rate of GSH transport. Chemical analysis revealed that intact GSH accounted for most of vesicle-associated radioactivity. The initial rate of transport followed saturation kinetics with respect to GSH concentration; an apparent Km of 0.33 mM and V of 1.47 nmol/mg protein in 20 s were calculated. These results indicate that transport of GSH across the canalicular membranes is a carrier-mediated process. Replacement of NaCl in the transport medium by KCl, LiCl or choline chloride had no effect on the transport activity of the vesicles. The rate of GSH uptake by the vesicles was enhanced by valinomycin-induced K+-diffusion potential (vesicle inside-positive) and was inhibited by probenecid, indicating that GSH transport across the canalicular membranes is electrogenic and involves the transfer of negative charge. The transport of GSH was inhibited by oxidized glutathione or S-benzyl-glutathione. This transport system in canalicular plasma membranes may function in biliary secretion of GSH and its derivatives which are synthesized in hepatocytes by oxidative processes or glutathione S-transferase. 相似文献
18.
The budding capacity ofBegonia rex leaf fragments is well known; that of undetached leaves has been shown by us only recently after treating the leaves with 6γγ DMAAP. Benzyladenine is as effective as 6γγ DMAAP in stimulating budding. Lower temperatures (17°, 22–12°, 12°) are also capable of inducing bud formation but only after a small cut has been made in a main vein of the undetached leaf. Root formation can also be provoked on undetached leaves which have a cut in the main leaf vein by higher temperatures (24–22°) or by an IAA treatment. Differences in the first stages of bud formation on leaf fragments and on undetached leaves are observed using histochemical and histoautoradiographic techniques. 相似文献
19.
E Ogier-Denis G Trugnan A T Tran C Sapin M Aubery P Codogno 《Comptes rendus de l'Académie des sciences. Série III, Sciences de la vie》1989,310(3):41-47
The expression of O- and N-glycan chains during the enterocytic differentiation of HT-29 cells was followed using L-[63H]-fucose and D-[6-3H]-glucosamine radiolabeling. Whatever the cell population, i.e., differentiated or undifferentiated, the incorporation of radiolabeled sugars into glycoproteins was similar. However, differences among these two cell populations were noted when the ratio [3H]-glucosamine/[3H]-galactosamine and the sensitivity of glycopeptides to mild alkaline treatment were followed. From these data, we could conclude that there is a shift in the high molecular weight glycopeptides during the differentiation of HT-29 cells meaning an increase of O-linked glycopeptides correlated with a decrease of N-linked forms. 相似文献
20.
Occurrence of Shiga-like toxin-producing Escherichia coli in retail fresh seafood, beef, lamb, pork, and poultry from grocery stores in Seattle, Washington. 总被引:5,自引:3,他引:2 下载免费PDF全文
M Samadpour J E Ongerth J Liston N Tran D Nguyen T S Whittam R A Wilson P I Tarr 《Applied microbiology》1994,60(3):1038-1040
Fresh meat, poultry, and seafood purchased from Seattle area grocery stores were investigated for the presence of Shiga-like toxin-producing Escherichia coli by using DNA probes for Shiga-like toxin (SLT) genes I and II. Of the 294 food samples tested, 17% had colonies with sequence homology to SLT I and/or SLT II genes. 相似文献