Showing papers by "University of Basel published in 1978"

Neutron diffraction studies on selectively deuterated phospholipid bilayers

Abstract: NEUTRON diffraction combined with the use of selectively deuterated lipids can provide detailed information on the molecular structure of membranes. Because of the large difference between the coherent scattering length of hydrogen ( −3.74 fermis) and deuterium (6.67 fermis) the deuterated membrane segments show up as intense peaks in the neutron density profile and can thus easily be located in the membrane1–3. We have applied this method to bilayer membranes of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), selectively deuterated at 12 different positions in the polar head group and the hydrocarbon chains. We report here that the mean position of the deuterated segments within the membrane can be determined in most cases to a precision of better than ±1 A. The average orientation of the phosphocholine group in the gel state as well as in the liquid crystalline state is almost parallel to the membrane surface. In the gel state the two hydrocarbon chains are out of step by about 1.8 A, and water penetrates up to the glycerol backbone of the lipid molecules.

A specific glycoprotein as the target site of adhesion blocking Fab in aggregating Dictyostelium cells

Abstract: During the acquisition of aggregation competence a new antigen appears on the surface of Dictyostelium cells. Univalent antibody fragments (Fab) against this antigen render the cells unable to form the specific type of cell adhesion which is characteristic of aggregating cells. This membrane constituent has been purified and identified as a concanavalin A-binding glycoprotein present at about 2 X 10(5) copies per cell.

Influence of cholesterol on the polar region of phosphatidylcholine and phosphatidylethanolamine bilayers.

Abstract: The structural changes in the polar head group region of unsonicated bilayer membranes of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine produced by addition of cholesterol have been determined using deuterium and phosphorus-31 NMR. Incorportion of up to 50 mol percent cholesterol produces little change in the phosphorus-31 chemical shielding anisotropies, compared with the values in pure bilayers above the phase transition temperatures, while some of the deuterium quadrupole splittings are reduced by almost a factor of two. Adjustment of the head group torsion angles by only a few degrees accounts for the observed spectral changes. Addition of cholesterol therefore has opposite effects on the hydrocarbon and polar regions of membranes: although cholesterol makes the hydrocarbon region gel-like, with an increased probability of trans conformations, the conformation of the polar head groups is very similar to that found in the liquid crystalline phase of pure phospholipid bilayers.

New factor released by cultured glioma cells supporting survival and growth of sensory neurones.

Abstract: PROCESS formation can be induced in clonal neuroblastoma cells by a macromolecule termed glial factor which has been identified in medium conditioned by cultured C-6 glioma cells1. Fibre outgrowth can also be elicited in cultures of peripheral sensory neurones2 by the well-characterised mouse sub-maxillary gland nerve growth factor (NGF)3, which has been shown to be a different entity from glial factor4. We report here that glioma-conditioned medium (GCM), the source of glial factor, can support both survival and fibre formation of isolated chick sensory neurones and that neither NGF nor glial factor are responsible for this effect.

Elastic Electron Scattering from He 3 and He 4 at High Momentum Transfer

Abstract: Experimental values of $^{3}\mathrm{He}$ ($^{4}\mathrm{He}$) elastic structure functions up to momentum transfer ${q}^{2}=4.0 (2.4)$ ${(\mathrm{G}\mathrm{e}\mathrm{V}/\mathit{c})}^{2}$ are presented. They are compared to calculations using three- and four-body wave functions and to asymptotic models.

Physical Evidence for the Asembly of A and B Chains of Human Placental Collagen in a Single Triple Helix

Abstract: Native collagen molecules containing A and B chains were isolated from pepsin-solubilised human chorionic and amniotic membrane extracts by fractional salt precipitation and DEAE-cellulose chromatography. They exhibited a circular dichroism spectrum, and a melting curve, characteristic for a triple-helical structure. Electron microscopical investigations of their segment-long-spacing crystallites revealed a molecule similar to those of the interstitial types I, II and III collagens. After denaturation, the A and B chains were separated by DEAE-cellulose chromatography and were consistently recovered in a ratio of 1:2. Renaturation experiments indicated that only the B chains are able to reform triple-helical molecules which are stable under conditions in vivo. The data support a molecular formula A(B)2 for the native collagen molecule.

Is sex determination in germ line and soma controlled by separate genetic mechanisms

Abstract: IN sexually dimorphic animals, the determination of sex is a major branch point in development. Although mutations producing sex reversal shed some light on the processes by which this determination is reached, many questions remain unanswered. Autosomal mutations such as polled in the goat1, sex reversed (Sxr)2 in the mouse and transformer (tra)3,7 in Drosophila cause chromosomal females to become phenotypic males, although they are sterile and their gonads agametic. The nature of this sterility suggests that the sex of germ cells in Drosophila, and perhaps in other species, might be determined by different sets of genes from those which operate in somatic tissue. Here we test this hypothesis by constructing mosaics in Drosophila where germ cells of one genotype are surrounded by soma of another. We find that the transformer mutation affects only the somatic sexual differentiation, and that it has no effect on the differentiation of germ cells. Thus we conclude that the agametic nature of transformed flies is due to the absence of functional male germ cells and at least with respect to tra, germ line and soma have separate sex-determining genes.

Is water critical to the formation of micelles in apolar media

Abstract: Photon correlation studies in H2O/sodium di-2-ethylhexylsulfosuccinate (AOT)/i-C8H18-systems, pertinent IR. investigations, and vapor pressure osmometric measurements with alkylated quaternary ammonium di-2-ethylhexylsulfosuccinates strongly suggest water to be pre-requistite for the micellization in apolar media.

fs(1)K10 , a germline-dependent female sterile mutation causing abnormal chorion morphology in Drosophila melanogaster

Abstract: Females homozygous for a newly isolated mutation induced by ethyl methane sulphonate,fs(1)K10, lay abnormally shaped eggs in which the dorsal appendages of the chorion are enlarged and fused ventrally. The eggs are usually not fertilized and development is never normal beyond the blastoderm stage. The mutant was mapped to the tip of the X-chromosome with a meiotic position of 1–0.5 and a cytological location between 2B17 and 3A3. Using germ line mosaics constructed by transplantation of pole cells, it was shown that the abnormal morphology and the sterility are obtained only when the germ line is homozygous for the mutant.

The Role of Cis-Trans Isomerization of Peptide Bonds in the Coil ⇄ Triple Helix Conversion of Collagen

Abstract: The collagen-like portion of a peptide which comprises the amino-terminal precursor-specific region of bovine type III procollagen, showed an unusually fast, concentration independent and fully reversible triple helix ⇄ coil transition. A set of three interchain disulfide bridges probably provides an effective nucleus for triple helix formation. Refolding occurred in two kinetic phases. The first one was not resolved (half time < 5 s) and the second one had a half time of about 90 s at 20 °C. When measurements were performed in phosphate buffer pH 7.0, the amplitudes of both phases were of comparable size. Activation energies for the rate constant of the slow phase ranging from 44kJ/mol at 40–30 °C to 70 kJ/mol at 15–5 °C were observed. In the presence of 4 M guanidine-HCI, essentially no fast phase was found and the activation energy was 97 ± 13 kJ/mol. The occurrence of two kinetic phases was explained by a model mechanism in which a stretch of helix between the nucleus and the first cis peptide bond is formed quickly. In the following slow phase, cis-trans isomerization at the junction of a triple helical to coiled region becomes rate-limiting. This interpretation was supported by temperature double-jump experiments. The reformation of cis peptide bonds, after a fast destruction of the triple helix, was paralleled by an increase in the amplitude of the slow phase. These findings qualitatively agree with results on the role of cis-trans isomerization in the folding of ribonuclease [Brandts, J. F., Halvorson, H. R. & Brennan, M. (1975) Biochemistry, 14, 4953–4963]. The observed curved Arrhenius plot was quantitatively explained by the normal activation energy of cis-trans isomerization (85 kJ/mol) and the contribution of the negative enthalpy of the pre-equilibria between partially helical species formed in the fast phase. Our data suggest that for long collagen molecules, the fast phase is negligible and cis-trans isomerization sets an upper limit for the rate of triple helix formation in vivo. At 37 °C the minimum time estimated for complete folding is in the range of minutes and comparable with the rate of biosynthesis. At low temperatures the slow rate of cis-trans isomerization may govern the intracellular formation of native collagen molecules to a greater extent, if no other mechanisms maintain the trans configuration of peptide bonds after release of the chains from the ribosomes.

Three Conformationally Distinct Domains in the Amino-Terminal Segment of Type III Procollagen and Its Rapid Triple Helix ⇄ Coil Transition

Abstract: Peptide Col 1–3 which comprises the whole amino-terminal precursor-specific region of bovine type III procollagen was isolated after digestion of type III procollagen with bacterial collagenase at 37 °C. Further treatment of Col 1–3 with collagenase at 55 °C released two non-collagenous segments, Col 1 and Col 2. Ultracentrifugal and circular dichroism (CD) studies indicated that Col 1–3 consists of three chain constituents associated to each other by both disulfide bridges and non-covalent bonds. The three interchain disulfide bridges are located in fragment Col 2, i.e. near the carboxyl end of the precursor-specific region. The CD spectrum indicates a random coil structure for peptide Col 2. Peptide Col 1 possesses five intrachain disulfide bridges and shows a CD spectrum indicating aperiodic structural elements and β structure. The non-covalent association of the peptide chains is due to a triple helix formed from a collagenous segment (Col 3) located between the Col 1 and Col 2 domains of the peptide. This triple helix has a melting temperature of 53 °C which decreases by about 20 °C when the peptide is reduced and S-carboxymethylated. The thermal transition of Col 1–3 is accounted for by a non-staggering zipper model without nucleation difficulty, using a standard enthalpy of triple helix formation of δHθ=− 35 kJ/mol tripeptide unit or by an all or none mechanism with δHθ=− 12.5 kJ/mol tripeptide units. Refolding of the helix from denatured Col 1–3 occurred in 100% yield with a rate constant of 8 × 10−3 S−1 at 20 °C. This is the highest rate constant yet observed for triple helix formation in vitro. Prior reduction of disulfide bonds prevented rapid helix formation.

A proton nuclear-magnetic-resonance study of self-stacking in purine and pyrimidine nucleosides and nucleotides.

Abstract: The concentration dependence of the chemical shifts of the protons H-2, H-8 and H-1′ of ATP4− and of Mg(ATP)2−, of all non-labile protons of adenosine, of H-5, H-6 and H-1′ of UTP, and of H-5, H-6, H-1′, and H-2′ of uridine have been measured. The results for the purine derivatives are consistent with the isodesmic model of indefinite non-cooperative stacking; for adenosine K= 15 ± 2 M−1, for ATP K= 1.3 ± 0.2 M−1 and for Mg(ATP)2−K= 3.6 ± 0.3 M−1. For the pyrimidines, uridine and UTP, stacking is much weaker and the stability constant could only be estimated; for uridine K⋜ 0.5 M−1, and for UTP K≈ 0.3 M−1.

The mitochondria-rich cell of frog skin as hormone-sensitive "shunt-path".

Abstract: Further investigations about the role of the mitochondria-rich cell (MR cell) in hormone-mediated transport regulation in the epithelium of frog skin brought the following results: Unlike toad bladder, in frog skin the spontaneous potential difference cannot be reversed when Na transport is blocked. A similar situation is obtained when, in addition to transport-blockade, one applies a chemical gradient for chloride to the epithelium. Under these conditions we found that in the intact preparation as well as in the separated epithelium: (i) the reversed current (RC) is linearly related to the number of MR cells; (ii) RC is mainly carried by a passive, transcellular chloride flux inwards and (iii) RC is sensitive to nor-adrenaline (10(-7) M). The beta-blocker propranolol abolishes this effect. We propose that the MR cells are the sites of transepithelial shunt-path and that this chloride flux is transcellular. As it is hormone sensitive, it could be an important regulatory instrument for the regulation of overall salt transport (internal shorting).

Nucleosome arcs and helices.

Abstract: Crystals and other regular arrangements of nucleosome cores have been obtained and analyzed in the electron microscope. Two types of regular structures have been studied in detail, the nucleosome arcs and cylinders. The latter are composed of concentric cylindrical layers of intertwined right-handed helices of nucleosome cores. These studies lead to the following conclusions and concepts. The overall structure of the nucleosome core is a short, wedge-shaped cylinder measuring about 110 by 110 by 60 angstroms. Nucleosome cores interact primarily between top and bottom planes. Nucleosome cores exhibit large conformational variability. A pivot allowing two degrees of rotational freedom is postulated in the region of the 70th base pair to account for this property of the nucleosome.

E. coli K-12 pel mutants, which block phage lambda DNA injection, coincide with ptsM, which determines a component of a sugar transport system.

Abstract: Escherichia coli pel - mutants inhibit the penetration of bacteriophage lambda DNA into the cell. Using P1 mediated cotransduction, we mapped pel - mutations between markers fadD and eda in the interval of minute 40 of the revised E. coli K-12 map. This places pel in the same region as genes kdgR and ptsM. Mutations in kdgR usually do not alter the Pel phenotype, and vice versa. In contrast, about 30% of ptsM - mutants are also pel -, and all pel - mutants isolated are ptsM -. These results suggest that pel and ptsM are one and the same gene. This interpretation would identify the bacterial product required for injection of phage λ DNA as a component of the phosphoenolpyruvate-dependent phosphotransferase system specific for mannose, glucosamine, glucose and fructose. However, the experimental results do not exclude an alternative explanation: that pel and ptsM identify two closely linked genes which would be simulataneously affected at high frequency by a particular mutational event.

An efficient method to produce specific anti-actin

Abstract: The production of affinity column purified (specific) anti-actin is described With the immunization scheme employed, all rabbits produced precipitating antibodies over several months, so that 30 mg specific anti-actin per rabbit could be isolated in 6 months The antibodies against native and detergent denatured smooth muscle actin are characterized by immunodiffusion tests, staining of the I-band of isolated myofibrils and stress fibers in tissue culture cells, using indirect immunofluorescence

Note on Time Evolution of Non-Markov Processes

Abstract: We discuss the question of the construction of a linear semigroup for the time evolution of the single-event probabilities of general non-Markov processes. It is shown that such a linear semigroup may not exist for all finite times. The consequences are sketched for the description of equilibrium and nonequilibrium systems. Further, the relationship with nonstationary Markov processes is investigated, and some confusion in recent works is cleared up using the example of free Brownian motion.