Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The rates of uridine-5-(3)H incorporation into RNA and the rates of uridine uptake into the acid-soluble pool during the cell cycle of V79 Chinese hamster cells were examined. Cells cultured on Eagle's minimal essential medium supplemented with fetal calf serum, lactalbumin hydrolysate, glutamine, and trypsin displayed rates of incorporation and uptake which increased only slightly during G(1) and accelerated sharply as DNA synthesis commenced. In contrast, cells cultured on minimal essential medium supplemented only with calf serum exhibited rates of incorporation and uptake which increased linearly through both G(1) and S. The transition from one pattern to the other can be induced within 24 hr and is completely reversible. The nonlinear pattern exhibited by cells grown on the supplemented fetal calf serum medium can also be overcome with high exogenous uridine concentrations. In the presence of 200 microM uridine, these cells display a linear pattern of increase in rates of uridine incorporation and uptake. It is concluded that at lower uridine concentrations the pattern of increase in the rate of uridine incorporation into RNA during the cell cycle for a given population of cells is dependent upon the rate of uridine entry into the cell, and that this pattern is not rigidly determined but can be modified by culture conditions.
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PMID:Induced changes in the rates of uridine- 3 H uptake and incorporation during the G 1 and S periods of synchronized Chinese hamster cells. 500 16

1. Treatment of Micrococcus lysodeikticus polynucleotide phosphorylase (nucleoside diphosphate-polynucleotide nucleotidyltransferase) with trypsin causes a preferential loss of its cytidine diphosphate and uridine diphosphate polymerization activities. 2. The phosphorolytic activity of the enzyme towards polycytidylic acid is unaffected in conditions in which the cytidine diphosphate-polymerization activity without added primer is virtually abolished. 3. The treated enzyme retains its altered pattern of activities when purified fivefold by gel filtration. 4. The effect on the cytidine diphosphate-polymerization activity is due, in part, to a large increase in primer requirement as a result of proteolysis, and is qualitatively independent of the state of purity of the polynucleotide phosphorylase. 5. The enzyme is protected from trypsin degradation by nucleic acids, polynucleotides and nucleoside disphosphates. 6. A similar, but less marked differential effect, is caused by alpha-chymotrypsin.
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PMID:The effect of trypsin digestion on the activities of polynucleotide phosphorylase. 605 26

The occurrence of insulin receptors and biological responses to insulin has been investigated in trypsin-dissociated fetal rat brain cells maintained in culture for 8 days. Binding of [125I]insulin to brain cells in culture was time- and pH-dependent and 85--90% specific. Porcine insulin competed for [125I]insulin binding in a dose-dependent manner. Unrelated polypeptides, including angiotensin II, glucagon, bovine growth hormone, and bovine prolactin did not compete for [125I]insulin binding. The half-life of [125I]insulin dissociation from receptors at 24 degrees C was 15 min and a plot of In[B/Bo] vs time suggested two dissociated rate constants of 2.7 X 10(-4) sec-1 and 5.0 X 10(-5) sec-1. Scatchard analysis of the binding data gave a curvilinear plot which may indicate negative cooperativity or the occurrence of both high affinity (Ka = 2 X 10(11) M-1) and low affinity (Ka = 4 X 10(10) M-1) sites. Of the estimated total of 4.9 X 10(4) binding sites per cell, 28--30% appear to be high affinity sites. Incubation of cultures with insulin caused a time- and dose-dependent stimulation of [3H]thymidine and [3H]uridine incorporation into TCA-precipitable material. Maximum stimulation of thymidine incorporation (2--5-fold) occurred 11 h after incubation with 167 nM insulin. The same concentration of insulin caused a 2.2-fold increase in [3H]uridine incorporation in 2 h. These results indicate that cells cultured from rat brain contain specific insulin receptors capable of mediating effects of insulin on macromolecular synthesis in the central nervous system.
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PMID:Binding of [125I]insulin to specific receptors and stimulation of nucleotide incorporation in cells cultured from rat brain. 615 64

Significant changes in the nucleus structure, complete suppression of the mitotic activity, markedly decreased synthesis of RNA (by 70--80 per cent according to incorporation of 3H-uridine) and decreased levels of DNA (by 40 per cent according to olivomycin binding) were observed in the fibroblasts cultivated in vitro due to exposure to actinoxanthine in an amount of 50 microgram/ml. The data indicate direct damaging effect of the drug on the cell chromatin. The above nuclear changes were also observed after a short-term exposure of the cells to the drug (up to 5 minutes). Still, they became evident only after the subsequent incubation of the cells in a pure culture medium for at least 15 minutes. No such changes in the nucleus structure were detected when after the 5-minute exposure to actinoxanthine the cells were exposed to trypsin for 3 minutes. When the time of exposure to actinoxanthine was longer (15 minutes and higher), trypsin suppressed the manifestation of the above nuclear changes. The two-stage mechanism of the damaging effect of actinoxanthine on the chromatin of the cells cultivated in vitro is discussed. The damaging effect of actinoxanthine on the cells begins from binding of the drug with the cell membrane. After that a short incubation period follows and then the characteristic changes in the nucleus structure appear.
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PMID:[Effect of the antitumor antibiotic actinoxanthine on cells cultured in vitro]. 617 13

In the presence of porcine submaxillary N-acetylgalactosaminyltransferase and uridine diphospho-N-acetyl-D-galactosamine, approx. 1.2-1.5 mol of N-acetylgalactosamine were transfered per mol of myelin basic protein. Tritium-labelled N-acetylgalactosamine-labelled basic protein was digested with trypsin and the peptides were separated by HPLC and the radioactivity measured. Most of the radioactivity was associated with three peptide peaks (I, II and III) containing 17, 69 and 6% of the total radioactivity, respectively. The remaining radioactivity was distributed amongst several peptides, each containing less than 2.5% of the total radioactivity. Glycosylation of the basic proteins isolated from human, bovine and guinea pig myelins showed that they were all equally good acceptors. In spite of differences in the peptide profiles of the basic proteins from different species, the distribution of radioactivity between the three peptide peaks was similar for all the species studied. The transfer of N-acetylgalactosamine to peptide II was much faster than to peptides I and III. The apparent Km values of the three peptides were within a narrow range of 0.52-0.63 mM, whereas the Vmax values were considerably different. The glycosylated peptide peaks (I, II and III) were separated by electrophoresis, the radioactivity measured, and amino acid compositions determined after hydrolysis. The major radioactive peptides of the human basic protein were identified with tryptic peptides containing the following sequences: (formula; see text)
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PMID:Identification of the major sites of enzymic glycosylation of myelin basic protein. 619 25

Human rotaviruses were capable of efficient multiplication in LLC-MK2 cells when the inoculum was pre-treated with trypsin, centrifuged on to the cell monolayer and the infected cells maintained in a medium containing trypsin. However, not all of the human rotavirus isolates used to infect cells resulted in efficient virus production. The ability of human isolates to multiply in cultured cells was studied by direct observation of virus in the electron microscope, by radioactive labelling with 3H-uridine of the newly synthesized virus and by electron microscopic examination of thin sectioned infected cells. With one of the specimens used (F-617) only 5 to 10% of the cells showed evidence of virus multiplication, with the great majority of the infected cells showing numerous complete (double-capsid) virus particles scattered in the cytoplasm. When cells were inoculated with another human specimen (SIB-I), infected cells were more abundant, reaching a maximum of 60%; however, a variety of particle types, probably representing different subviral structures or different steps of rotavirus morphogenesis, were commonly observed. The presence of these aberrant or incomplete virus structures may represent a manifestation of the defectiveness of this virus and may explain the difficulties encountered in its serial passage.
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PMID:Multiplication of human rotavirus in cultured cells: an electron microscopic study. 624 81

Protoplasts of Bacillus subtilis W23 readily synthesized ribitol teichoic acid from nucleotide precursors in the surrounding medium. With cytidine diphosphate-ribitol they made poly(ribitol phosphate), presumably attached to lipoteichoic acid carrier; when cytidine diphosphate-glycerol and uridine diphosphate-N-acetylglucosamine were also present a 10-fold increase in the rate of polymer synthesis occurred, and the product contained both the main chain and the linkage unit. Synthesis was inhibited by trypsin or p-chloromercuribenzenesulfonate in the medium, and we concluded that it occurred at the outer surface of the membrane. During synthesis, which was also achieved readily by whole cells after a brief period of wall lysis, the cytidine phosphate portion of the nucleotide precursors did not pass through the membrane. No evidence could be obtained for a transphosphorylation mechanism for the translocation process. It is suggested that reaction with exogenous substrates was due to temporary exposure of a protein component of the enzyme complex at the outer surface of the membrane during the normal biosynthetic cycle.
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PMID:Synthesis of teichoic acid by Bacillus subtilis protoplasts. 627 28

1. Nucleoside transport by fetal erythrocytes from nucleoside-permeable and nucleoside-impermeable type new-born lambs and by reticulocytes from adult sheep was compared with that of mature erythrocytes from adult sheep of the two phenotypes.2. Fetal cells and reticulocytes transported [U-(14)C]uridine rapidly, with little difference between cells from the two types of sheep. Transport occurred by a saturable uptake mechanism with similar properties to that present in mature cells from adult nucleoside-permeable type animals, except for an approximately 100-fold higher V(max).3. This increased translocation capacity was associated with increased numbers of high-affinity [(3)H]nitrobenzylthioinosine binding sites ( approximately 2000-3000 sites/cell compared with approximately 20 sites/cell for mature nucleoside-permeable sheep erythrocytes).4. The calculated transport capacity for each nucleoside translocation site is therefore similar in all cell types (140-180 molecules/site. s at 25 degrees C, assuming that each transport site binds a single molecule of inhibitor). These values compare favourably with turnover estimates for the nucleoside transporter from human and pig erythrocytes.5. Loss of nucleoside transport activity after birth closely paralleled loss of [(3)H]nitrobenzylthioinosine binding sites and the progressive loss of fetal cells from the circulation. Similarly, reticulocyte maturation in vitro was also associated with rapid loss of both nucleoside transport capacity and inhibitor binding activity.6. p-Chloromercuriphenylsulphonate and trypsin had no effect on [(3)H]nitrobenzylthioinosine binding to intact fetal cells. In contrast, both agents markedly inhibited binding to isolated ;ghosts' where both sides of the cell membrane were accessible to reagent. p-Chloromercuriphenylsulphonate inhibition was markedly reduced in the presence of uridine, and reversed by addition of dithiothreitol.7. We conclude that nucleoside transport changes during ontogeny and reticulocyte maturation in the sheep as well as species differences in nucleoside transport capacity are regulated by variations in the numbers of functional transport sites per cell rather than by changes in the activity of a constant number of sites. It is also likely that the nucleoside carrier exhibits chemical asymmetry.8. A simple molecular model of the erythrocyte nucleoside transporter consistent with these and other known properties of the carrier is proposed.
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PMID:Nucleoside translocation in sheep reticulocytes and fetal erythrocytes: a proposed model for the nucleoside transporter. 628 22

Evidence is presented for the first time that a human candidate calicivirus (HCV) replicates in human embryo kidney cells when trypsin is incorporated in the culture medium. The virus multiplies in the presence of actinomycin D and radiolabelling experiments with [3H]uridine indicate that it has an RNA genome. These observations provide further support for the view that HCV should be tentatively classified as a member of the Caliciviridae .
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PMID:Propagation of human candidate calicivirus in cell culture. 672 90

A fraction purified from acetic acid extracts of porcine hypothalami was found to contain significant antimitogenic activity when tested in normal and neoplastic cell lines. Addition of this hypothalamic material (1-100 micrograms/ml) to culture media significantly inhibited [3H]thymidine incorporation into cellular DNA in several cell lines. Amino acid incorporation into pituitary proteins and uridine incorporation into RNA were also significantly reduced by this factor(s). Addition to the culture media of this hypothalamic material at 5 micrograms/ml and 50 micrograms/ml per day decreased by 17% and 36%, respectively, cell numbers of 3T6 fibroblast cell cultures. Time-response curves showed that the inhibition of [3H]thymidine incorporation into DNA in 3T6 fibroblast cells begins within 2 hr after adding this fraction to the culture medium. The inhibitory action cannot be explained by a direct cytotoxic effect since 3T6 cells labeled with 51Cr and incubated for 6 hr in the presence of this hypothalamic fraction fail to show an increase in the release of 51Cr into the medium as compared with controls. Incubation with trypsin and chymotrypsin completely abolished the antimitogenic activity of this material and pepsin decreased it. This strongly suggests that the antimitogenic activity exhibited by this fraction is due to a polypeptide(s). These observations provide evidence for the presence in the mammalian hypothalamus of an antimitogenic peptide(s) that may be involved in the regulation of cell proliferation.
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PMID:Inhibition of cell growth by a hypothalamic peptide. 675 25


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