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Biomedical subjects

G Maass

Publications and source records attributed to G Maass.

At least 91 records · Page 5Linked to original sources
PubMed1986

[Vegetative hyperthermia: a thermoregulation disorder or a variant from the norm?].

A diagnosis of low-grade idiopathic fever was made in 85 women and 15 men, examined for subfebrile temperature of unknown origin, after organic disease had been excluded. Compared with 100 healthy control subjects these patients had inadequate movement-dependent temperature elevations with a usually pronounced discrepancy between the rectal temperature, predominantly more than 38 degrees C, and a normal or only slightly raised axillary temperature recorded after bodily movement. Other characteristics were that the elevated temperature was uninfluenced by antipyretic drugs. Almost all patients initially complained about general lassitude as well as frequently about atypical functional complaints and psychological symptoms. These observations indicate that low-grade fever is a functional syndrome due to a harmless faulty regulation of body temperature. In addition to a special constitutional reactivity, psychological factors and possibly previous febrile infections are likely to be involved as precipitating causes.

Adolescent↗
PubMed1986

Polypeptide sequences involved in the cleavage of DNA by the restriction endonuclease EcoRI.

We have prepared a variety of fragments of the restriction endonuclease EcoRI by partial or total CNBr or acid cleavage of the protein. These fragments were isolated by preparative polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. They were analyzed in a qualitative manner for phosphodiesterase activity. Antibodies against these fragments were elicited in rats and tested for binding to native EcoRI in an enzyme-linked immunoassay. We conclude from these experiments that the DNA binding site of EcoRI is located in the COOH-terminal half of the molecule, close to and probably comprising amino acid residues 137 to 157. This conclusion is reinforced by the observation that this sequence shows homology to the sequences of the recognition helix of other gene-regulatory proteins.

Amino Acid Sequence↗
PubMed1986

Conformation of the DNA undecamer 5'd(A-A-G-T-G-T-G-A-T-A-T) bound to the single-stranded DNA binding protein of Escherichia coli. A time-dependent transferred nuclear Overhauser enhancement study.

A time-dependent transferred nuclear Overhauser enhancement study of the conformation of the single-stranded DNA 11mer 5'd(A-A-G-T-G-T-G-A-T-A-T) bound to the single-stranded DNA binding protein of Escherichia coli (SSB) is presented. It is shown that the conformation of the bound 11mer is that of a right-handed B-type helix similar to that of the free 11mer. The observation of internucleotide transferred nuclear Overhauser enhancements for every base step excludes the possibility of intercalation by aromatic protein residues. In addition, it is shown that the effective correlation time of the bases (80 ns) corresponds to that of a complex of molecular weight approximately 170,000, containing two SSB tetramers. The sugars, on the other hand, exhibit a shorter effective correlation time (40 ns), indicating the presence of internal motion. This suggests that the bases are anchored to the protein surface, possibly by hydrophobic interactions, whereas the sugar-phosphate groups are directed outwards towards the solvent.

Bacterial Proteins↗
PubMed1986

Effect of 4-quinolones and novobiocin on calf thymus DNA polymerase alpha primase complex, topoisomerases I and II, and growth of mammalian lymphoblasts.

The influence of ciprofloxacin, nalidixic acid, norfloxacin, novobiocin, and ofloxacin on elements of eucaryotic DNA replication was investigated in vitro. Each of the 4-quinolones, when present in amounts of more than 100 micrograms/ml, reversibly inhibited the DNA synthesis performed by the 95 DNA polymerase alpha primase complex from calf thymus. Novobiocin at 500 micrograms/ml or at higher concentrations irreversibly inactivated DNA polymerase alpha primase complex. The accuracy of in vitro DNA synthesis in the absence of repair mechanisms was determined from amber-revertant assays with phi X174am16(+) DNA as template. The antimicrobial agents did not significantly increase the frequencies of base pairing mismatches during the course of replication, indicating that the basal mutation rate is not affected by novobiocin and the 4-quinolones. The Ki values of 50% inhibition of DNA topoisomerases from calf thymus by ciprofloxacin, norfloxacin, novobiocin, nalidixic acid, and ofloxacin were 300, 400, 1,000 or more, 1,000 or more, and 1,500 or more micrograms/ml, respectively, in the case of topoisomerase I, and the Ki values were 150, 300, 500, 1,000, and 1,300 micrograms/ml, respectively, in the case of topoisomerase II. The procaryotic topoisomerase II is approximately 100-fold more sensitive to inhibition by ciprofloxacin, norfloxacin, and ofloxacin than is its eucaryotic counterpart. Growth curves of lymphoblasts were recorded in the presence of ofloxacin and ciprofloxacin. Neither 1 nor 10 micrograms of ciprofloxacin or of ofloxacin per ml affected cell proliferation. Ofloxacin and ciprofloxacin at 100 micrograms/ml inhibited cell growth; 1,000 micrograms/ml led to cell death. No correlation exists between the antimicrobial and cytotoxic activities of the 4-quinolones.

Cell Division↗
PubMed1985

Linear diffusion of restriction endonucleases on DNA.

We have investigated the dependence of the rate of cleavage of DNA by EcoRI, HindIII, and BamHI on the chain length of the substrate. In order to keep the influence of flanking sequences and of nonspecific binding identical for all substrates we have carried out all experiments with the same plasmid DNA which had been digested previously with a variety of different restriction enzymes to give a set of substrates of different lengths. Our results show that depending on the buffer conditions long substrates are cleaved faster than small ones. We interpret these findings to mean that under certain conditions a linear diffusion of the enzymes on the DNA is involved in localizing the recognition sites. For EcoRI the mean diffusion length is approximately 1000 base pairs at 1 mM MgC12 which can be shown by diffusion theory to correspond to a linear diffusion coefficient of 5 X 10(-10) cm2 s-1. At 10 mM MgCl2 the linear diffusion of EcoRI is negligible and does not lead to a significant enhancement of the rate of site localization. In the presence of nonsaturating amounts of one of the prokaryotic histone-like protein Hu (NS 2) small and large DNA substrate are cleaved with identical rate by EcoRI indicating that other proteins bound to the DNA constitute a barrier across which linear diffusion cannot take place. We conclude that linear diffusion, albeit detectable under certain conditions in vitro, probably is of little importance for the process of site localization in vivo.

Binding Sites↗
PubMed1985

Detection of human rotavirus by nucleic acid analysis in comparison to enzyme-linked immunoassay and electron microscopy.

The results of RNA analysis for the detection of rotavirus were compared with those of a standard enzyme-linked immunosorbent assay (ELISA) and electron microscopy using 212 faecal specimens obtained from 200 children with gastroenteritis. Rotavirus was extracted directly from faecal specimens and RNA segments were made visible by polyacrylamide gel electrophoresis using a silver staining technique. Of the 212 faecal specimens 137 were found to be positive in ELISA, 125 in RNA analysis and 121 in both methods. Forty-nine of the 212 specimens were also investigated by electron microscopy. Thirty-five were positive when examined by electron microscopy, 37 were positive in ELISA and 33 in RNA analysis. RNA analysis of 119 faecal samples in outbreaks and sporadic cases of rotavirus infection yielded 42 different rotavirus electrophoretypes. The results indicated that no one method was sufficient to detect all positive specimens and that RNA analysis is useful in epidemiological studies.

Electrophoresis, Polyacrylamide Gel↗
PubMed1984

Escherichia coli single-stranded DNA binding protein is mobile on DNA: 1H NMR study of its interaction with oligo- and polynucleotides.

The interaction of the Escherichia coli single-stranded DNA binding protein (SSB) with oligo- and poly-nucleotides has been studied by 270-MHz 1H NMR spectroscopy and fast kinetic techniques. d(pT)8 and poly(dT) were used to study noncooperative and cooperative binding, respectively. The H6, H1', and CH3 resonances of d(pT)8 are high-field shifted by less than 0.05 ppm, and H8 and H2 of poly(dA) are low-field shifted upon complexation. The protein resonances remain virtually unshifted. The small shifts upon complexation provide no evidence for extensive stacking interactions between the nucleotide bases and aromatic amino acid side chains of SSB. The d(pT)8 and poly(dA) signals are broadened to about 30 Hz whereas the resonances of poly(dT) are broadened beyond detection upon stoichiometric complexation. Continuous broadening of all poly(dT) signals even at a 10-fold excess of poly(dT) indicates fast exchange of SSB between different binding sites. Dissociation and reassociation rates determined from stopped-flow experiments are too slow by at least 2 orders of magnitude to account for the experimental line widths. Therefore, we conclude that SSB translocates without dissociation from the DNA template. A model for the translocation is outlined. It is based on partial dissociation of octamer sections of poly(dT) from the complex with a rate constant as previously published for the dissociation of d(pT)8 from SSB.

DNA-Binding Proteins↗
PubMed1984

Anthracenyl crown ethers and cryptands as fluorescent probes for solid-phase transitions of phosphatidylcholines: syntheses and phospholipid membrane studies.

Three structurally related crown compounds and cryptands have been synthesized that differ by the number and linkage of coronand units and anthracene moieties. The interaction of the fluorescent dyes with sonicated dimyristoylphosphatidylcholine (DMPC) vesicles is characterized by the relative quantum yields, uptake kinetics, binding curves, lifetimes, fluorescence titrations with water- and lipid-soluble quenching agents, fluorescence anisotropy, and equilibrium cooling curves. The most lipophilic compound II, which displays a similar quantum yield as the parent fluorophore 9,10-dimethylanthracene, shows a nearly equal distribution between solid and fluid lipid and is located at the bilayer surface. The least lipophilic compound IV is excluded from the hydrocarbon phase. The anthracenophane cryptand III preferentially partitions into solid-phase lecithins with the highest affinity for the phases L epsilon and L beta. The binding constant to DMPC amounts to (5.4 +/- 1.3) X 10(2) M-1 at 0 degrees C. From fluorescence quenching titrations it is concluded that the average position of the anthracenoyl dye III discontinuously shifts during the gel to liquid crystalline transition from the glycerol backbone to the choline head group. Electron microscopy and NMR experiments revealed that the anthracenophane induces in the liquid crystalline phase the fusion of small unilamellar DMPC vesicles to unilamellar medium-sized vesicles and macrovesicles, which subsequently fuse at the transition temperature to large multilamellar coacervates. Due to its large change of fluorescence intensity, the anthracenophane cryptand is a very sensitive probe for the detection of the pretransition of symmetrically substituted and of the subtransition of asymmetrically substituted phosphatidylcholines.(ABSTRACT TRUNCATED AT 250 WORDS)

Anthracenes↗
PubMed1984

Kinetics of the subtransition of asymmetrically substituted phosphatidylcholines.

The thermodynamics and kinetics of the subtransition L epsilon----P beta of sonicated unilamellar vesicles of 1-myristoyl-2-stearoylphosphatidylcholine (1M-2S-PC) and of 1-stearoyl-2-myristoylphosphatidylcholine (1S-2M-PC) were studied by equilibrium cooling curves and temperature-jump relaxation spectrometry with an anthracenophane cryptand as a mobile fluorescent probe. The unilamellar vesicles exhibit the midpoint temperature TsII of the subtransition about 10 degrees C below the respective main transition. The kinetics of the subtransition in the time range between 10(-4) and 10(3) s is characterized by a cooperative relaxation process in the range of milliseconds and a further noncooperative process in the range of seconds. The slow process is assigned to the rearrangement of lattice defects. The fast process is evaluated in terms of a cyclic reaction scheme that consists of two pathways for the biomolecular association of probe and vesicle coupled with the conformational change of the lipid matrix during the subtransition. The analysis reveals that the fast process comprises the nucleation and growth of cluster. The cooperative lattice transformation of the subtransition follows a first-order rate law. The rate constants at TsII are 70 s-1 for 1S-2M-PC and 170 s-1 for 1M-2S-PC. Since the plots of the relaxation time vs. the degree of transition are in accordance with the predictions of the linear Ising model, it is concluded that clusters are propagated anisotropically in a linear fashion; e.g., fluidlike P beta conformations grow along the ripple.

Kinetics↗
PubMed1984

The influence of sequences adjacent to the recognition site on the cleavage of oligodeoxynucleotides by the EcoRI endonuclease.

We have investigated the influence of the nucleotide sequence adjacent to the recognition site on the rate of cleavage of DNA by the restriction endonuclease EcoRI. For this purpose two decadeoxynucleotides, d(G-G-G-A-A-T-T-C-T-T) (Ia) and d(A-A-G-A-A-T-T-C-C-C) (Ib) were synthesized. The duplex Ia X Ib is cleaved by EcoRI preferentially in the dA-rich strand (approximately 10 times over the dG-rich strand). The individual nucleotides Ia and Ib are also cleaved by EcoRI, Ib at a higher rate than Ia and both at a lower rate than Ia X Ib. The temperature dependence of the reaction rate shows that only double-stranded oligodeoxynucleotides are substrates for the EcoRI endonuclease. We have, furthermore, synthesized oligomers of d(G-G-A-A-T-T-C-C), which contain two, three and four EcoRI sites, respectively. These oligodeoxynucleotides are preferentially cleaved at the sites next to the 5' end, where the recognition site is only flanked by one dG X dC base pair, in contrast to the other sites which are flanked by three such pairs. These data indicate that sequences adjacent to the recognition site influence the rate of cleavage: dA X dT base pairs enhance and dG X dC base pairs slow down the hydrolytic activity of the EcoRI endonuclease.

Base Composition↗
PubMed1983

Does the specific recognition of DNA by the restriction endonuclease EcoRI involve a linear diffusion step? Investigation of the processivity of the EcoRI endonuclease.

The time course of the EcoRI endonuclease catalysed cleavage of three substrates, two plasmid DNAs and one oligonucleotide, each with two EcoRI sites, was measured. The two plasmid DNAs with the EcoRI sites 318 and 96 base pairs apart are cut in a distributive fashion, while the oligonucleotide with the EcoRI sites 8 base pairs apart is cut in a partially processive manner. It is concluded that a linear diffusion of the EcoRI endonuclease on its substrate across long stretches of DNA is not likely to be operative during the recognition process. Microscopic dissociation-reassociation processes, however, increase the probability of the enzyme to attack further sites located in the immediate vicinity of a given site.

Base Composition↗
PubMed1983

Isolation and identification of enteric adenoviruses.

Thirty-four out of 64 faecal samples with adenovirus particles, as seen by electron microscopy, were found to contain adenovirus 40 or 41 by direct isolation and neutralization in Chang's conjunctival cells, mostly within one week. (Ad40 and 41 candidate viruses are serologically related.) 6 other adenovirus species were isolated; 6 samples gave equivocal results, and 18 were negative. A genus-specific ELISA with an antihexon coat yielded positive results in 40 out of 55 samples; the test failed to identify adenovirus antigen in 10 out of 17 specimens, which were found negative by culture. All of them were negative by immunfluorescence of inoculated Chang cell cultures. Hence the failures are probably due to insufficient amount of virus in the samples. The predominance of only two adenovirus species associated with gastroenteritis in children and the ease of cultivating and identifying them should help to elucidate their etiological significance.

Adenoviridae Infections↗
PubMed1983

Problems of live virus vaccine-associated poliomyelitis a paralytic case with isolation of all three poliovirus types.

The case of a 4-month-old girl is described who developed a paralytic polio-like syndrome 3 weeks after oral polio vaccination (OPV). All three poliovirus types could be isolated (9 days after onset of disease polio type 2, and 33 days after onset of disease types 1 and 3, respectively). In order to classify these isolates as Sabin (vaccine)-like (SL) or non-Sabin-like (non-SL), several markers were tested in three laboratories [intratypic serodifferentiation, reproductive capacity at supraoptimal temperature (RTC), Al(OH)3 gel elution assay, and oligonucleotide mapping]. The results of the marker determinations were not uniform, but--summarizing all data--it seems plausible to associate the disease with the OPV. The significance of marker determinations in proving a vaccine-induced poliomyelitis is discussed in the light of this clinical case. Some comments are made on poliovirus vaccination policy in developed countries.

Antibodies, Viral↗
PubMed1983

Coronavirus-like particles as aetiological agents of acute non-bacterial gastroenteritis in humans.

The morphology of coronavirus-like particles found in faecal extracts of neonates and infants with acute gastroenteritis show similarities with the structure of human coronaviruses. These coronavirus-like particles can be detected in faeces of sick neonates and infants in higher frequency than in healthy children of the same age, their association with this clinical syndrome is discussed. The significance of enteric coronavirus-like particles in acute gastroenteritis of adults is questionable since the particles can be found in faeces of diseased persons and of healthy adults with equal frequency.

Acute Disease↗
PubMed1982

Two identical subunits of the EcoRI restriction endonuclease Co-operate in the binding and cleavage of the palindromic substrate.

The cleavage of radioactively labelled double-stranded d(G-G-A-A-T-T-C-C) was studied in single turnover experiments with substrate and enzyme both being in the micromolar range. The reaction rate was found to increase with enzyme concentration until a ratio of one tetrameric enzyme to two double-stranded substrates was reached, further increase of the enzyme concentration then leads to a sharp decline of the reaction rate. These findings are interpreted in the following manner. (a) Two subunits of the EcoRI endonuclease co-operate in binding and possibly also in cleaving the palindromic substrate. (b) The enzymatic action of the EcoRI endonuclease is inhibited by excess enzyme, possibly due to unspecific binding of the enzyme-substrate complex. The self-inhibition of EcoRI endonuclease has also been observed with macromolecular substrates.

Binding, Competitive↗
PubMed1981

The effect of several nucleic acid binding drugs on the cleavage of d(GGAATTCC) and pBR 322 by the Eco RI restriction endonuclease.

The endonucleolytic action of the EcoRI restriction enzyme on the double-stranded oligonucleotide d(GGAATTCC) and the supercoiled plasmid DNA pBR 233 is inhibited by actinomycin D, ethidium bromide, proflavin, distamycin A and netropsin. Half-maximal inhibition is observed at around 100 microM concentrations for the intercalating drugs, and around 0.1 to 1 microM concentrations for netropsin and distamycin A. The inhibitory activity of these drugs can be correlated with their affinity to the oligonucleotide and the plasmid DNA. Since at high concentrations of the drugs a complete inhibition is observed, it is concluded that the effect of the drugs on the stereochemistry of the EcoRI site is such that recognition is excluded.

Circular Dichroism↗
PubMed1981

Escherichia coli single-strand deoxyribonucleic acid binding protein: stability, specificity, and kinetics of complexes with oligonucleotides and deoxyribonucleic acid.

The complex formation between the single-strand DNA binding protein (ssB protein) from Escherichia coli and oligonucleotides and single-stranded DNA has been studied by using fluorescence titrations, ultracentrifugation measurements, and fast kinetic techniques. Determination of the stoichiometries of oligo(dT)--ssB complexes shows that each of the four subunits of the ssB protein represents a binding site for an oligonucleotide about eight residues long. Occupation of all four binding sites with oligo(dT) or poly(dT) leads to 80% quenching of the intrinsic protein fluorescence. The binding sites are nearly equivalent and independent. For d(pT)16, the intrinsic binding constant is 6 X 10(5) M-1, and for d(pT)30-40, which is long enough to extend continuously over the ssB tetramer, the binding constant is higher than 5 X 10(8) M-1. Oligoadenylates bind about 2 orders of magnitude weaker than the corresponding oligo(dT) species. The binding of oligo(dT) is very weakly dependent on ionic strength, in contrast to the oligo(dA)--ssB complex formation. For d(pT)8, d(pT)16, and d(pT)30-40, the complex formation can be described by a simple one-step reaction. The strength of the interaction is mainly expressed in the rate constant of dissociation. In the cooperative complexes with poly(dT) or poly(dA), all four binding sites on the ssB tetramer are also occupied. It is concluded that single-stranded DNA is coiled around the ssB molecule. Fluorescence melting experiments of the complexes show that the conformation of the single-stranded DNA has a strong influence on the stability of the complexes.

Binding Sites↗