Search PubMed⌕ Search

Biomedical subjects

M Bachmann

Publications and source records attributed to M Bachmann.

At least 145 records · Page 8Linked to original sources
PubMed1986

Purification and characterization of the major nucleoside triphosphatase from rat liver nuclear envelopes.

Nuclear envelopes contain a nucleoside triphosphatase. Hydrolysis of ATP or GTP by this enzyme parallels energy-dependent efflux of poly(A)-containing mRNA from nuclei in vitro. Nucleoside triphosphatase has been purified from highly purified preparations of nuclear envelopes from rat liver by three successive affinity steps. The essentially homogeneous enzyme has an apparent molecular weight of 40,000 as checked by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and displays a rather broad substrate specificity. ATP and GTP are hydrolyzed at nearly equal rates, whereas UTP and CTP are only half as active as substrates. For optimal activity, a one-to-one ratio of a divalent cation (Mg2+, Mn2+, or Ca2+) and the nucleoside triphosphate substrate, an alkaline pH and a temperature of 34 degrees C are required. In contrast to the enzyme associated with nuclear envelopes which is stimulated by synthetic poly(A) and the poly(A) segment of the natural poly(A)-containing mRNA, homogeneous nucleoside triphosphatase is unable to be modulated by this polynucleotide species.

Adenosine Triphosphate↗
PubMed1986

Association of La and Ro antigens with intracellular structures in HEp-2 carcinoma cells.

Monoclonal antibodies were raised against homogeneous Ro and La antigens, two proteins associated with Ro and La ribonucleoproteins (RNPs). The specificity of the monoclonal antibodies was proven by immunoblot analysis and by immunoprecipitation. The anti-Ro antibody reacted with a Mr 95,000 protein in a mouse lymphoma cell extract and with a Mr 60,000 polypeptide in extracts from human spleen. The anti-La antibody recognized a Mr 50,000 polypeptide in the mouse L5178y cell extract. The two monoclonal antibodies precipitated RNPs that contained the typical RNA species of Ro or La RNPs. The localization of Ro and La antigen was performed by direct immunofluorescence microscopy. It was found that the anti-Ro antibody reacted with a fibrous network that behaves like cytokeratin, one of the intermediate filament systems. The anti-La antibody reacted with nuclear structures that gave a speckled-type pattern.

Animals↗
PubMed1986

Identification and further characterization of the specific cell binding fragment from sponge aggregation factor.

Monoclonal antibodies (McAbs) were raised against the aggregation factor (AF) from the marine sponge Geodia cydonium. Two clones were identified that secrete McAbs against the cell binding protein of the AF complex. Fab fragments of McAbs: 5D2-D11 completely abolished the activity of the AF to form secondary aggregates from single cells. The McAbs were determined to react with the AF in vitro; this interaction was prevented by addition of the aggregation receptor, isolated and purified from the same species. After dissociation of the AF by sodium dodecyl sulfate and 2-mercaptoethanol, followed by electrophoretical fractionation, a 47-kD protein was identified by immunoblotting which interacted with the McAbs: 5D2-D11. During this dissociation procedure, the sunburst structure of the AF was destroyed. In a second approach, the 47-kD protein was isolated by immunoprecipitation; 12 molecules of this protein species were calculated to be associated with the intact AF particle. The 47-kD AF fragment bound to dissociated Geodia cells with a high affinity (Ka of 7 X 10(8) M-1) even in the absence of Ca++ ions; the number of binding sites was approximately 4 X 10(6)/cell. This interaction was prevented by addition of the aggregation receptor to the 47-kD protein in the homologous cell system. Moreover, it was established that this binding occurs species-specifically. The 47-kD fragment of the AF was localized only extracellularly by indirect immunofluorescence staining in cryostat slices. These data suggest that the 47-kD protein is the cell binding molecule of the AF from Geodia.

Animals↗
PubMed1986

Occurrence of novel small RNAs with concomitant inhibition of host cellular U small nuclear RNA synthesis in Vero cells infected with herpes simplex virus type 1.

In eukaryotic cells, small nuclear and small cytoplasmic RNAs (sn- or scRNAs) are associated with distinct proteins, forming ribonucleoproteins (snRNPs or scRNPs). In the present study we analysed the protein composition as well as the small RNA pattern in non-infected and herpes simplex virus type 1 (HSV)-infected Vero cells. We found that concomitantly with the shut-off of host cell mRNA synthesis, synthesis of U-snRNAs was stopped. Due to their stability, however, U-snRNAs were still present in cells 36 h after HSV infection. Besides these RNAs, two novel small RNAs which we termed HVR1 and HVR2 were detected in infected cells. On the basis of their relative mobilities in urea gels, the apparent chain lengths of these newly synthesized RNAs were determined to be 255 and 154 nucleotides respectively. The small RNA-binding proteins Sm, RNP, Ro and La were found to increase up to 15-fold after HSV infection. The data presented suggest that new, virus-coded small RNPs are synthesized which might play a role in the maturation and regulation of HSV-coded RNA transcripts.

Animals↗
PubMed1986

Purification and characterization of the Ro and La antigens. Modulation of their binding affinities to poly(U) by phosphorylation and the presence of ATP.

Both the La and the Ro antigen (the latter for the first time) were purified to apparent homogeneity. Ro was found to be a 94 (90)-kDa and La a 50-kDa polypeptide. Both antigens bind to RNA with a high preference for poly(U). The binding hierarchy is U much greater than G greater than A greater than C for La and U much greater than C greater than G greater than A for Ro. Only 15% of the total amount of La or 21% of that of Ro, present in the L5178y cell extract, is able to bind to poly(U), indicating the existence of RNA binding and nonbinding subclasses of La and Ro. The purified antigens were used for the isolation of monospecific antibodies. These antibodies were specific for their respective antigen and did not cross-react. Both the Ro and the La antigen are phosphorylated in vitro by the cytoplasmic protein kinase CII, whereas the nuclear protein kinases NI and NII are unable to phosphorylate the antigens. After phosphorylation or in the presence of ATP the binding affinity of both antigens to poly(U) strongly decreases. The phosphorylation reaction together with the immunoprecipitation by the monospecific antibodies represents a highly sensitive and specific assay which was used during purification and characterization of the Ro and La antigen.

Adenosine Triphosphate↗
PubMed1985

[The occurrence of goitrogenic substances in milk. 1. Release of goitrin in the milk of cows fed on rapeseed extract cakes].

A total of six cows, divided into 3 groups, were fed various amounts of rape cake containing 6 g of goitrin/kg over a period of 7 days. The cows were milked twice a day and the goitrin content of the heated milk samples were determined by a HPLC-method within 2 h. When rape cake was fed at 0.39, 1.9 and 3.9% resp. of the total feed this resulted in medium goitrin values of 37, 163 and 707 micrograms/l milk. These values correspond to a transfer of about 0.1% of the original progoitrin content in the feed. 12 h after the last rape feeding the amount of goitrin in the milk was below the detection limit of 7 ppb. The toxicological significance of these findings are dicussed.

Animal Feed↗
PubMed1985

Simultaneous measurement of serotonin and propranolol pulmonary extraction in patients after extracorporeal circulation and surgery.

To evaluate the use of markers of pulmonary endothelial cell function in humans, we simultaneously measured the pulmonary extraction of serotonin (SER) and propranolol (PROP) in patients before and after extracorporeal circulation (ECC) for coronary bypass surgery. No change was seen in SER extraction ratio after anesthesia, ECC, and surgery. Twenty-four hours after ECC and surgery, PROP extraction ratio was decreased by 11%. It remained low for as long as 5 days thereafter. A similar drop in PROP extraction was found in 6 patients 24 h after ilioaortic bypass surgery, without ECC (from 81 to 66%, p less than 0.01). Treatment of 11 other patients with a 30-min period of continuous positive airway pressure (CPAP), 24 to 48 h after ECC, resulted in a significant increase in PROP extraction ratio from 61 to 67% (p less than 0.01). Measurement of PROP extraction appears to be more sensitive than that of SER to lung changes related to postoperative atelectasis.

Adult↗
PubMed1985

Pulmonary extraction of serotonin and propranolol in patients with adult respiratory distress syndrome.

Because injury to the pulmonary vascular endothelium is associated with the development of the adult respiratory distress syndrome (ARDS), we assessed the metabolic function of pulmonary endothelial cells by the measurements of the first-pass pulmonary extraction of [14C]serotonin and [3H]propranolol in 15 patients with ARDS and 15 patients at risk for developing ARDS. Serotonin extraction ratio was lower in patients with ARDS (0.85 +/- 0.10, mean +/- SD) than in patients at risk (0.91 +/- 0.04) (p less than 0.025), and both values were significantly reduced (p less than 0.005) when compared with a control group value (0.97 +/- 0.01). The decrease in serotonin extraction was correlated with the severity of ARDS (r = -0.67) (p less than 0.001) and with pulmonary function changes over time. Propranolol extraction ratio was decreased in patients at risk (0.66 +/- 0.11) (p less than 0.005) but not in patients with ARDS (0.75 +/- 0.11), when compared with those in the control group (0.81 +/- 0.03). Low values in patients at risk were restored to normal by continuous positive airway pressure breathing. We conclude that pulmonary extraction of serotonin, an index of pulmonary endothelial cell function, correlates with the severity of ARDS.

Adult↗
PubMed1984

12 S small nuclear ribonucleoprotein-associated acidic-and pyrimidine-specific endoribonuclease from calf thymus and L5178y cells.

12 S ribonucleoprotein (RNP) particles were separated from a 45 S RNP complex (Bachmann, M., Zahn, R. K. and Müller, W. E. G. (1983) J. Biol. Chem. 258, 7033-7040) isolated from calf thymus and L5178y cells. The particles were determined to be associated with an acidic endoribonuclease (pI 4.1; pH optimum 6.2). the enzyme requires Mg2+ and is sensitively inhibited by higher NaCl concentrations. The nuclease specifically degrades poly(U) and poly(C) in an endonucleolytic manner; the end-products are 3'-UMP (85%) and 2',3'-cyclic UMP (12%). Poly(A) strongly inhibits the pI 4.1 endoribonuclease activity. The Michaelis constant (for poly(U)) was determined as 82 microM and the maximal reaction velocity was 0.54 mumol/microgram per h. The endoribonuclease is distinguished from the known pyrimidine-specific ribonucleases (pancreatic ribonuclease and endoribonuclease VII) by further criteria, e.g., resistance to thiol reagents, inhibition by EDTA, Mg2+ requirement, pI and pH optimum. Using the techniques of counterimmunoelectrophoresis and immunoaffinity column chromatography it was shown that the pI 4.1 endoribonuclease-associated 12 S RNP particles display antigenicity to anti-Sm and anti-(U1)-RNP antibodies. An RNA component, isolated from the 12 S-45 S hypercomplex, was identified as U1-snRNA.

Animals↗
PubMed1984

The role of protein phosphokinase and protein phosphatase during the nuclear envelope nucleoside triphosphatase reaction.

The activities of nuclear envelope-associated protein phosphokinase and protein phosphatase were determined in nuclear ghosts from liver and oviduct of quails. The protein kinase was found to be inhibited by poly(A) by 75%. During the kinase reaction proteins with molecular weights of 106 000 and 64 000 were phosphorylated. The phosphoprotein phosphatase from liver was stimulated to 190% by poly(A), whereas only a slight enhancing effect by this polymer was determined with the oviduct enzyme (to 125%). Comparative determinations of the nuclear ghost-associated enzyme activities revealed the following values (in nmol Pi/min per 10(8) ghosts); oviduct: phosphokinase, 0.015; phosphatase, 0.004 and nucleoside triphosphatase, 39.4; and liver: phosphokinase, 0.044; phosphatase, 0.012 and nucleoside triphosphatase, 11.7. These data indicate that phosphorylation/dephosphorylation proceeds independently of the nucleoside triphosphatase cycle. This assumption is supported by analytical results revealing that no marked dephosphorylation occurs after poly(A) binding to the nuclear envelope. Moreover, stoichiometrical data showed a nearly 1:1 molar ratio between ATP-binding and phosphorylation of nuclear envelope protein. From these findings a new model for the nucleoside triphosphatase-mediated poly(A)(+)mRNA efflux from nuclei is deducted, proposing phosphokinase and phosphatase only to modulate the affinity of the 'carrier structure' for poly(A) (+)mRNA, but not to constitute the nucleoside triphosphatase.

Animals↗
PubMed1984

Base-specific ribonucleases potentially involved in heterogeneous nuclear RNA processing and poly(A) metabolism.

Polyadenylation and splicing of heterogeneous nuclear RNA, two crucial steps in mRNA processing, are apparently enzymatically mediated processes. This contribution summarizes the properties and the presumed functions of the known poly(A) catabolic enzymes (endoribonuclease IV and V, 2',3'- exoribonuclease ) as well as those of the pyrimidine-specific endoribonucleases associated with snRNP -hnRNP complexes (endoribonuclease VII, acidic pI 4.1 endoribonuclease and poly(U)-specific U1 snRNP -nuclease).

Animals↗
PubMed1984

Goitrin--a nitrosatable constitutent of plant foodstuffs.

N-nitrosamides are known as direct-acting carcinogens at the site of their formation; they do not need any metabolic activation in vivo. The conditions leading to their formation in the stomach, and also their genotoxicity, have been thoroughly studied with some model compounds. Several reports link this type of compound to the induction of gastric cancer in human. However, only limited data are presently available about possible precursors of N-nitrosamides in foods. In the present study we found that goitrin --a naturally occurring compound in cruciferous vegetables and rape--could be easily nitrosated by treatment with nitrite under stomach conditions, yielding with loss of sulfur the N-nitroso- oxazolidone 4 (fig.). This product has a mutagenicity pattern and potency similar to that of N-nitroso-N-methyl-N'- nitroguanidine (MNNG) in the Ames Salmonella/mammalian microsome test.

Biotransformation↗
PubMed1984

Age-dependent changes of nuclear envelope protein phosphokinase and protein phosphatase activities. Significance for altered nucleo-cytoplasmic mRNA translocation during development.

Nuclear envelopes are associated with a protein phosphokinase and a phosphoprotein phosphatase, whose activities are modulated by poly(A) in an opposite manner. The activities of these enzymes were determined in nuclear ghosts from liver and oviduct of quails of different age and of different hormone status. Under optimal conditions, kinase activity was found to increase in immature animals 8-fold in response to diethylstilbestrol; co-administration of progesterone had no marked effect on enzyme activity. After the initial burst, the activity of the enzyme increased only slightly during ageing. Two proteins present in nuclear ghosts of Mr 64 000 and of Mr 106 000 are phosphorylated during the kinase reaction; both the relative and the absolute extents of phosphate incorporation into these proteins alter drastically during ageing or hormone treatment of immature animals. Like the kinase activity, the activity of protein phosphatase increased in immature animals markedly in response to hormone treatment. Thereafter the activity remained constant in liver while in oviducts the phosphatase activity dropped to 30% in both mature and old animals.

Aging↗
PubMed1984

Effect of intravenous midazolam on breathing pattern and chest wall mechanics in human.

Breathing pattern, thoracoabdominal motion, and separate end-expiratory positions of the rib cage and abdomen were measured noninvasively in eight healthy subjects before and after intravenous administration of either placebo or midazolam, a short-acting benzodiazepine. Compared with placebo, midazolam produced a significant (P less than 0.01) decrease in mean inspiratory flow of 29% from preinjection values, resulting in a 39% reduction in tidal volume (VT). This ventilatory depression was partly compensated by a 35% decrease in expiratory time producing an increase in respiratory rate (+39%). The fall in VT was almost entirely (91%) mediated by a reduction of the abdominal contribution to tidal breathing while sparing rib cage motion. This fact contrasts with the effects of inhalational anesthetics or morphine, which preferentially depress rib cage expansion, indicating that thoracoabdominal motion may selectively be depressed by different pharmacological agents. In addition, continuous recording of end-expiratory levels showed a significant transient fall in the rib cage's end-tidal position 2 min after midazolam administration associated with the occurrence of central apneas.

Abdominal Muscles↗
PubMed1984

Synergistic effect of peplomycin in combination with bleomycin on L5178y mouse lymphoma cells in vivo.

Studying the treatment of NMRI mice with ip injections of bleomycin (BLM) for 5 days we found an approximate LD50 of 35 mg/kg; the toxicity of peplomycin (PEP) was slightly higher (LD50: approximately 25 mg/kg). The effect of the two drugs on growth of L5178y mouse lymphoma cells in NMRI mice was examined. BLM alone caused at a concentration of 2.5 mg/kg an almost complete inhibition of tumor cell growth; the same effect was determined with 1 mg PEP/kg. At these concentrations the drugs caused an increase of the survival time of 110% (BLM) or 104% (PEP). Given in combination, one-sixth of the optimal doses yielded an 100% increase of the median survival time. These results indicate a significant synergistic activity of the PEP-BLM combination on L5178y cell growth in vivo (FIC index: 0.34).

Animals↗
PubMed1983

Association of a polyuridylate-specific endoribonuclease with small nuclear ribonucleo-proteins which had been isolated by affinity chromatography using antibodies from a patient with systemic lupus erythematosus.

Immunoglobulins, containing antibodies against U1-snRNP, have been prepared from a patient with systemic lupus erythematosus. After coupling these antibodies to a Sepharose matrix, U-snRNPs have been isolated and purified from rat liver nuclei by use of immunoaffinity chromatography. The resulting RNPs had the typical protein pattern of U-sn RNPs and a sedimentation coefficient of 12 S. The U-snRNP preparation was associated with an endoribonuclease which required Mg2+ for optimal activity. The enzyme, with an pH optimum of 6.2, degraded only poly(U). Other single-stranded polyribo- and polydeoxyribonucleotides, tRNA, as well as double-stranded RNA and DNA were not digested. The products of a terminal digestion are (U)6-12 with 3'-OH and 5'-P termini. The possible involvement of this endoribonuclease in the splicing of hnRNA is discussed.

Animals↗
PubMed1983

Purification and properties of a novel pyrimidine-specific endoribonuclease termed endoribonuclease VII from calf thymus that is modulated by polyadenylate.

Endoribonuclease VII, a novel endoribonuclease from calf thymus, was identified and purified by us. The purified enzyme has Mr = 74,000; its homogeneity was checked by analysis in polyacrylamide gels (both in the presence and in the absence of sodium dodecyl sulfate). The nuclease cleaves poly(U) and poly(C) while other single-stranded homopolyribo- as well as polydeoxyribonucleotides are not degraded; poly(A,C) is hydrolyzed to a smaller extent, while poly(U) X poly(A) is not degraded at all. Poly(A) modulates the poly(U)-degrading activity; at a molar ratio of approximately 1 [poly(A)]:10 [poly(U)], a more than 100% stimulation of the enzyme activity was achieved, while at lower ratios an almost complete inhibition of the enzyme activity resulted. Binding studies revealed that endoribonuclease VII has a marked affinity for poly(A) and poly(U). During hydrolysis, oligo(U)12 fragments with 3'-OH and 5'-P termini are formed. The basic enzyme (pI = 8.5) has its activity optimum at pH 7.2, requiring neither monovalent nor divalent cations; the enzyme is not inhibited by thiol group reagents. Several lines of evidence suggesting a role of endoribonuclease VII in mRNA processing are presented.

Animals↗