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G Rapp

Publications and source records attributed to G Rapp.

At least 37 records · Page 2Linked to original sources
PubMed1998

X-Ray Diffraction Study of the Lamellar-Hexagonal Phase Transition in Phospholipid/Surfactant Mixtures.

The lamellar-to-hexagonal phase transition of a phospholipid/ surfactant mixed system of 1-palmitoyl-2-oleoyl-sn-glycero-3phosphocholine (POPC) and oligo(ethylene oxide)dodecyl ethers of type C12H25O(CH2CH2O)2H(C12E2) in molar surfactant/phospholipid ratio (RS/L) of 2 at low hydration driven by temperature has been studied by X-ray diffraction. The Lbeta-HII phase transition is a reversible two-state process showing hysteresis at fast temperature scan rates. The obtained hexagonal phase exhibits a temperature dependent structural change. The numbers of bound water molecules per composite particle (WS+L) absorbed in the lamellar and hexagonal phases are nearly the same, changing from WS+L = 5.0 to 4.7 during the phase transition. The fluidity of the alkyl chains on increasing the temperature and the close packing of the hydrophilic molecular parts are the driving parameters of the lamellar-to-hexagonal transformation. Copyright 1998 Academic Press.

Journal Article↗
PubMed1998

Structural characterization of the pressure-denatured state and unfolding/refolding kinetics of staphylococcal nuclease by synchrotron small-angle X-ray scattering and Fourier-transform infrared spectroscopy.

The pressure-induced unfolding of wild-type staphylococcal nuclease (Snase WT) was studied using synchrotron X-ray small-angle scattering (SAXS) and Fourier-transform infrared (FT-IR) spectroscopy, which monitor changes in the tertiary and secondary structural properties of the protein upon pressurization. The experimental results reveal that application of high-pressure up to 3 kbar leads to an approximate twofold increase of the radius of gyration Rg of the native protein (Rg approximately 17 A) and a large broadening of the pair-distance-distribution function, indicating a transition from a globular to an ellipsoidal or extended chain structure. Analysis of the FT-IR amide I' spectral components reveals that the pressure-induced denaturation process sets in at 1.5 kbar at 25 degrees C and is accompanied by an increase in disordered and turn structures while the content of beta-sheets and alpha-helices drastically decreases. The pressure-induced denatured state above 3 kbar retains nonetheless some degree of beta-like secondary structure and the molecule cannot be described as a fully extended random coil. Temperature-induced denaturation involves a further unfolding of the protein molecule which is indicated by a larger Rg value and significantly lower fractional intensities of IR-bands associated with secondary-structure elements. In addition, we have carried out pressure-jump kinetics studies of the secondary-structural evolution and the degree of compactness in the folding/unfolding reactions of Snase. The effect of pressure on the kinetics arises from a larger positive activation volume for folding than for unfolding, and leads to a significant slowing down of the folding rate with increasing pressure. Moreover, the system becomes two-state under pressure. These properties make it ideal for probing multiple order parameters in order to compare the kinetics of changes in secondary structure by pressure-jump FT-IR and chain collapse by pressure-jump SAXS. After a pressure jump from 1 bar to 2.4 kbar at 20 degrees C, the radius of gyration increases in a first-order manner from 17 A to 22.4 A over a timescale of approximately 30 minutes. The increase in Rg value is caused by the formation of an extended (ellipsoidal) structure as indicated by the corresponding pair-distance-distribution function. Pressure-jump FT-IR studies reveal that the reversible first order changes in beta-sheet, alpha-helical and random structure occur on the same slow timescale as that observed for the scattering curves and for fluorescence. These studies indicate that the changes in secondary structure and chain compactness in the folding/unfolding reactions of Snase are probably dependent upon the same rate-limiting step as changes in tertiary structure.

Kinetics↗
PubMed1998

Fibrillar structure and mechanical properties of collagen.

Collagen type I is among the most important stress-carrying protein structures in mammals. Despite their importance for the outstanding mechanical properties of this tissue, there is still a lack of understanding of the processes that lead to the specific shape of the stress-strain curve of collagen. Recent in situ synchrotron X-ray scattering experiments suggest that several different processes could dominate depending on the amount of strain. While at small strains there is a straightening of kinks in the collagen structure, first at the fibrillar then at the molecular level, higher strains lead to molecular gliding within the fibrils and ultimately to a disruption of the fibril structure. Moreover, it was observed that the strain within collagen fibrils is always considerably smaller than in the whole tendon. This phenomenon is still very poorly understood but points toward the existence of additional gliding processes occurring at the interfibrillar level.

Animals↗
PubMed1998

Studies on the 14.5 nm meridional X-ray diffraction reflection during length changes of intact frog muscle fibres.

The intensity of the 14.5 nm meridional reflection (M3) from activated skeletal muscle fibres was studied in both single fibres and fibre bundles during the imposition of length changes. During shortening at small load, the intensity of the reflection decreased within 2 ms to less than 20% of isometric intensity, then recovered partially during the remainder of the shortening. When shortening was terminated, recovery of intensity was delayed. Small shortening steps (0.5% fibre length) produced a fall in M3 intensity (IM3) delayed by ca. 250 microseconds compared to the fall in tension. For larger step releases (1% fibre length), the fall in IM3 was not delayed. The fall in IM3 could be almost completely reversed by a subsequent restretch applied within 1.5 ms. Beyond 10 ms after the initial release, the restretch caused a further fall in intensity. A rapid step stretch (0.5% fibre length) also caused a fall in IM3 without delay, which was partially reversed by a release applied within 10 ms. A second small release applied 3 ms (or less) after the first caused a second fall in M3 intensity, but without delay and with faster time course. Small amplitude sinusoidal length oscillations (0.15-0.2% sarcomere; 1 kHz) caused a sinusoidal change in M3 intensity, which was 180 degrees out of phase with the force oscillations, and lacked distortion during its release phase.

Animals↗
PubMed1998

X-ray diffraction studies of the cross-bridge intermediate states.

Two dimensional x-ray diffraction was obtained from skinned rabbit psoas muscle fibers. The goal is to correlate structures of the cross-bridge population with various intermediate states in the cross-bridge cycle by using nucleotides and their analogs. It was found that in a relaxed muscle in ATP containing solutions, cross-bridges are distributed in three populations in equilibrium: those detached from actin and ordered on the myosin helix, those that are detached and disordered, and those weakly attached to actin in random orientations. The distribution among the three populations is highly dependent on temperature. Those that are detached and yet ordered in a helical structure surrounding the myosin backbone are very likely in the M.ADP.Pi state, supporting an earlier suggestion by Wray (1987). It was also found that the attached cross-bridges with bound MgADP are structurally distinct from those without nucleotide, in agreement with one of our earlier findings by osmotic compression (Xu et al., 1993). Another finding of interest is that the analog AMP-PNP was found to be an ATP analog, rather than an ADP analog as it has been reported previously by many research groups.

Adenosine Triphosphate↗
PubMed1998

Evidence for charge-controlled conformational changes in the photocycle of bacteriorhodopsin.

The existence of two different M-state structures in the photocycle of the bacteriorhodopsin mutant ASP38ARG was proved. At pH 6.7 (0 to -6 degreesC) a spectroscopic M intermediate (M1) that does not differ significantly in its tertiary structure from the light-adapted ground state accumulates under illumination. At pH > 9 another state (M2), characterized by additional pronounced changes in the Fourier transform infrared difference spectrum in the region of the amide I and II bands, accumulates. The M2 intermediate trapped at pH 9.6 displays the same changes in the x-ray diffraction intensities under continuous illumination as previously described for x-ray experiments with the mutant ASP96ASN. These observations indicate that in this mutant the altered charge distribution at neutral pH controls the tertiary structural changes that seem to be necessary for proton translocation.

Bacteriorhodopsins↗
PubMed1998

Accelerated formation of cubic phases in phosphatidylethanolamine dispersions.

By means of x-ray diffraction we show that several sodium salts and the disaccharides sucrose and trehalose strongly accelerate the formation of cubic phases in phosphatidylethanolamine (PE) dispersions upon temperature cycling through the lamellar liquid crystalline-inverted hexagonal (Lalpha-HII) phase transition. Ethylene glycol does not have such an effect. The degree of acceleration increases with the solute concentration. Such an acceleration has been observed for dielaidoyl PE (DEPE), dihexadecyl PE, and dipalmitoyl PE. It was investigated in detail for DEPE dispersions. For DEPE (10 wt% of lipid) aqueous dispersions at 1 M solute concentration, 10-50 temperature cycles typically result in complete conversion of the Lalpha phase into cubic phase. Most efficient is temperature cycling executed by laser flash T-jumps. In that case the conversion completes within 10-15 cycles. However, the cubic phases produced by laser T-jumps are less ordered in comparison to the rather regular cubic structures produced by linear, uniform temperature cycling at 10 degrees C/min. Temperature cycles at scan rates of 1-3 degrees C/min also induce the rapid formation of cubic phases. All solutes used induce the formation of Im3m (Q229) cubic phase in 10 wt% DEPE dispersions. The initial Im3m phases appearing during the first temperature cycles have larger lattice parameters that relax to smaller values with continuation of the cycling after the disappearance of the Lalpha phase. A cooperative Im3m --> Pn3m transition takes place at approximately 85 degrees C and transforms the Im3m phase into a mixture of coexisting Pn3m (Q224) and Im3m phases. The Im3m/Pn3m lattice parameter ratio is 1. 28, as could be expected from a representation of the Im3m and Pn3m phases with the primitive and diamond infinite periodic minimal surfaces, respectively. At higher DEPE contents ( approximately 30 wt%), cubic phase formation is hindered after 20-30 temperature cycles. The conversion does not go through, but reaches a stage with coexisting Ia3d (Q230) and Lalpha phases. Upon heating, the Ia3d phase cooperatively transforms into a mixture of, presumably, Im3m and Pn3m phases at about the temperature of the Lalpha-HII transition. This transformation is readily reversible with the temperature. The lattice parameters of the DEPE cubic phases are temperature-insensitive in the Lalpha temperature range and decrease with the temperature in the range of the HII phase.

Crystallization↗
PubMed1998

Volume changes of the myosin lattice resulting from repetitive stimulation of single muscle fibers.

Single muscle fibers at 1 degreesC were subjected to brief tetani (20 Hz) at intervals of between 20 s and 300 s over a period of up to 2 h. A band lattice spacing increased during this period at a rate inversely dependent on the rest interval between tetani. Spacing increased rapidly during the first 10 tetani at a rate equivalent to the production of 0.04 mOsmol.liter-1 of osmolyte per contraction, then continued to expand at a much slower rate. For short rest intervals, where lattice expansion was largest, spacing increased to a limiting value between 46 and 47 nm (sarcomere length 2.2 micrometer), corresponding to accumulation of 30 mOsmol.liter-1 of osmolytes, where it remained constant until repetitive stimulation was terminated. At this limiting spacing, force was reduced by up to 30%. The effect of lattice swelling on the lattice compression that accompanies isometric force recovery from unloaded shortening was to increase the compression, similar to that observed in hypotonic media at a similar spacing. During recovery from repetitive stimulation, spacing recompressed to its original value with a half-time of 15-30 min. These findings suggest that mechanical activity produces an increase in osmotic pressure within the cell as a result of product accumulation from cross-bridge and sarcoplasmic reticulum ATPases and glycolysis.

Animals↗
PubMed1997

Low amounts of PEG-lipid induce cubic phase in phosphatidylethanolamine dispersions.

By using time-resolved X-ray diffraction we demonstrate that low amounts (5-10 mol%) of a phospholipid with two saturated hydrocarbon acyl chains 14 carbon atoms long and PEG550 chain covalently attached to its phosphoethanolamine polar head group, DMPE(PEG550), induce spontaneous formation of a cubic phase with lattice constant 20.5 nm (cubic aspect #8, space group Im3m) in aqueous dispersions of dielaidoylphosphatidylethanolamine (DEPE). This phase displays a highly resolved X-ray diffraction pattern with 17 low-angle reflections. The cubic phase was found to intrude in the temperature range between the lamellar liquid crystalline (L(alpha)) phase and the inverted hexagonal phase (H(II)) known to form in pure DEPE/water dispersions. A higher DMPE(PEG550) amount of 20 mol% was found to eliminate the non-lamellar phases in the temperature scale up to 100 degrees C. DMPE grafted with PEG5000 only shifts the L(alpha)-H(II) transition of DEPE to higher temperatures but does not promote formation of cubic phase. These findings indicate that, consistent with their bulky head groups, the PEG-lipids decrease the tendency for negative interfacial mean curvature of the DEPE bilayers.

Liposomes↗
PubMed1997

The tertiary structural changes in bacteriorhodopsin occur between M states: X-ray diffraction and Fourier transform infrared spectroscopy.

The tertiary structural changes occurring during the photocycle of bacteriorhodopsin (BR) are assigned by X-ray diffraction to distinct M states, M1 and M2. Purple membranes (PM) of the mutant Asp96Asn at 15, 57, 75 and 100% relative humidity (r.h.) were studied in a parallel X-ray diffraction and Fourier transform infrared (FTIR) spectroscopic investigation. Light-dependent conformational changes of BR-Asp96Asn are observed at high hydration levels (100 and 75% r.h.) but not in partially dehydrated samples (57 and 15% r.h.). The FTIR spectra of continuously illuminated samples at low and high hydration, despite some differences, are characteristic of the M intermediate. The changes in diffraction patterns of samples in the M2 state are of the same magnitude as those of wild-type samples trapped with GuaHCl in the M(G) state. Additional large changes in the amide bands of the FTIR spectra occur between M2 and M(G). This suggests, that the tertiary structural changes between M1 and M2 are responsible for the switch opening the cytoplasmic half-channel of BR for reprotonation to complete the catalytic cycle. These tertiary structural changes seem to be triggered by a charge redistribution which might be a common feature of retinal proteins also in signal transduction.

Amino Acid Sequence↗
PubMed1997

Enthalpy is a proper criterion for comparability of monolayer and bilayer studies: isobaric temperature scanning measurements on glycolipid monolayers.

The thermotropic behaviour of glycolipid monolayers has been studied by isobaric temperature scanning measurements to elucidate conditions under which monolayers exhibit thermodynamic and structural properties comparable to those observed in bilayers. A selection of synthetic, stereochemically pure, glyceroglycolipids with identical, ether-linked alkyl chains of 12, 14, or 16 CH2-groups has been investigated. The head groups of the glycolipids consisted of glucose, galactose, maltose, lactose or maltotriose moieties with beta-configuration of the glycosidic bond. These glycolipids were chosen to permit a quantitative characterization of three effects, (i) the role of the length of the aliphatic chains, (ii) the influence of the size of the head group, and (iii) the influence of the stereochemistry of the sugar moieties on the structure and stability of the monolayers. To probe the effects of stereochemical alterations in the glycerol moiety 2,3-O-ditetradecyl-1-O-beta-D-glucosyl-sn-glycerol (14-2,3-Glc) was compared with 1,2-O-ditetradecyl-3-O-beta-D-glucosyl-sn-glycerol (14-1,2-Glc). It has been shown that in general several features of bilayers can be obtained from monolayer studies with reasonable accuracy, provided the proper parameters are chosen. The monolayer is stabilized by elongation of the aliphatic chains of the lipids and destabilized when the monosaccharide read groups is replaced by a di-, or trisaccharide, in a similar manner as in the bilayer. The stabilizing effect that has been observed in bilayer studies, when galactose instead of glucose is introduced as head group, has also been established in the monolayer studies. This stabilizing effect is even retained in the lipids having disaccharide head groups. On the basis of these monolayer studies in connection with WAXS and SAXS measurements on multilamellar systems, we suggest that identity of transition enthalpies of the chain melting L beta-L alpha transition is an appropriate criterion for estimating molecular areas and area changes of bilayers from monolayer measurements and vice versa. However, estimates of transition temperatures are poor using the enthalpy criterion. If identity of transition temperature is introduced as criterion, glycolipid monolayers must be compressed to about 43 +/- 3 mN m-1. Under these conditions the agreement between the calculated enthalpies and structural properties of monolayers and multilayers is poor. As a general conclusion it can be emphasized that for monolayer and bilayer systems of glycolipids there exists no such parameter as a universal pressure or a universal temperature that automatically renders monolayer data identical to bilayer data. Depending on which property (transition temperatures, transition enthalpies, lateral areas and transitional area changes) one wants to extrapolate from monolayer to bilayer different lateral pressures have to be applied.

Calorimetry, Differential Scanning↗
PubMed1997

A new molecular model for collagen elasticity based on synchrotron X-ray scattering evidence.

Collagen is the most abundant structural protein in vertebrates. The specific shape of its stress-strain curve is crucial for the function of a number of organs. Although the macroscopic mechanical behavior of collagen is well known, there is still no explanation of the elastic process at the supramolecular level. We have performed in situ synchrotron x-ray scattering experiments, which show that the amount of lateral molecular order increases upon stretching of collagen fibers. In strain cycling experiments the relation between strain and diffuse equatorial scattering was found to be linear in the "heel" region of the stress-strain curve. A new molecular model for collagen elasticity is proposed, which, based on the existence of thermally activated molecular kinks, reproduces this linearity and gives a simple explanation for the form of the stress-strain curve of collagen.

Animals↗
PubMed1996

New phases induced by sucrose in saturated phosphatidylethanolamines: an expanded lamellar gel phase and a cubic phase.

A new lamellar gel phase (L beta *) with expanded lamellar period was found at low temperatures in dihexadecylphosphatidylethanolamine (DHPE) and dipalmitoylphosphatidylethanolamine (DPPE) dispersions in concentrated sucrose solutions (1-2.4 M). It forms via a cooperative, relatively broad transition upon cooling of the L beta gel phase of these lipids. According to the X-ray data, the transformation between L beta and L beta * is reversible, with a temperature hysteresis of 6-10 degrees C and a transition width of about 10 degrees C. No specific volume changes and a very small heat absorption of about 0.05 kcal/mol accompany this transition. The L beta *-L beta transition temperature strongly depends on the disaccharide concentration. From a value of about 10 degrees C below the melting transition of DHPE, it drops by 25 degrees C with decrease of sucrose concentration from 2.4 M to 1 M. The low-temperature gel phase L beta * has a repeat spacing by 8-10 A larger than that of the L beta gel phase and a single symmetric 4.2 A wide-angle peak. It has been observed in 1, 1.25, 1.5 and 2.4 M solutions of sucrose, but not in 0.5 M of sucrose. The data clearly indicate that the expanded lamellar period of the L beta * phase results from a cooperative, reversible with the temperature, increase of the interlamellar space of the L beta gel phase. Other sugars (trehalose, maltose, fructose, glucose) induce similar expanded low-temperature gel phases in DHPE and DPPE. The L beta * phase is osmotically insensitive. Its lamellar period does not depend on the sucrose concentration, while the lattice spacings of the L alpha, L beta and HII phases decrease linearly with increase of sucrose concentration. Another notable sugar effect is the induction of a cubic phase in these lipids. It forms during the reverse HII-L alpha transition and coexists with the L alpha phase in the whole temperature range between the HII and L beta phases. The cubic phase has only been observed at sucrose concentrations of I M and above. In accordance with previous data, sucrose suppresses the L alpha phase in both lipids and brings about a direct L beta-HII phase transition in DHPE. A raid, reversible gel-subgel transformation takes place at 17 degrees C in both DPPE and DHPE. Its properties do not depend on the sucrose concentration. The observed new effects of disaccharides on the properties of lipid dispersions might be relevant to their action as natural protectants.

Calorimetry, Differential Scanning↗
PubMed1995

Temperature- and pressure-dependent phase behavior of monoacylglycerides monoolein and monoelaidin.

We used x-ray and neutron diffraction to study the temperature- and pressure-dependent structure and phase behavior of the monoacylglycerides 1-monoelaidin (ME) and 1-monoolein (MO) in excess water. The monoacylglycerides were chosen for investigation of their phase behavior because they exhibit mesomorphic phases with one-, two-, and three-dimensional periodicity, such as lamellar, an inverted hexagonal and bicontinuous cubic phases, in a rather easily accessible temperature and pressure range. We studied the structure, stability, and transformations of the different phases over a wide temperature and pressure range, explored the epitaxial relations that exist between different phases, and established a relationship between the chemical structure of the lipid molecules and their phase behavior. For both systems, a temperature-pressure phase diagram has been determined in the temperature range from 0 to 100 degrees C at pressures from ambient up to 1400 bar, and drastic differences in phase behavior are found for the two systems. In MO-water dispersions, the cubic phase Pn3m extends over a large phase field in the T,p-plane. At temperatures above 95 degrees C, the inverted hexagonal phase is found. In the lower temperature region, a crystalline lamellar phase is induced at higher pressures. The phases found in ME-water include the lamellar crystalline Lc phase, the L beta gel phase, the L alpha liquid-crystalline phase, and two cubic phases belonging to the crystallographic space groups Im3m and Pn3m. In addition, the existence of metastable phases has been exploited. Between coexisting metastable cubic structures, a metric relationship has been found that is predicted theoretically on the basis of the curvature elastic energy approximation only.

Biophysical Phenomena↗
PubMed1994

Lattice spacing changes accompanying isometric tension development in intact single muscle fibers.

The myosin lattice spacing of single intact muscle fibers of the frog, Rana temporaria, was studied in Ringer's solution (standard osmolarity 230 mOsm) and hyper- and hypotonic salines (1.4 and 0.8 times standard osmolarity respectively) in the relaxed state, during "fixed end" tetani, and during shortening, using synchrotron radiation. At standard tonicity, a tetanus was associated with an initial brief lattice expansion (and a small amount of sarcomere shortening), followed by a slow compression (unaccompanied by sarcomere length changes). In hypertonic saline (myosin lattice compressed by 8.1%), these spacing changes were suppressed, in hypotonic saline (lattice spacing increased by 7.5%), they were enhanced. During unloaded shortening of activated fibers, a rapid lattice expansion occurred at all tonicities, but became larger as tonicity was reduced. This expansion was caused in part by the change in length of the preparation, but also by a recoil of a stressed radial compliance associated with axial force. The lattice spacing during unloaded shortening was equal to or occasionally greater than predicted for a relaxed fiber at that sarcomere length, indicating that the lattice compression associated with activation is rapidly reversed upon loss of axial force. Lattice recompression occurred upon termination of shortening under standard and hypotonic conditions, but was almost absent under hypertonic conditions. These observations indicate that axial cross-bridge tension is associated with a compressive radial force in intact muscle fibers at full overlap; however, this radial force exhibits a much greater sensitivity to lattice spacing than does the axial force.

Animals↗
PubMed1992

Construction of a plasmid containing the complete coding region of human elongation factor 2.

A plasmid pUChEF-2 containing the coding sequence as well as the complete 3'-untranslated region (3'UTR) of human EF-2 mRNA was constructed. The plasmid construct was assembled from a cDNA insert of pHGR81 (Rapp et al., (1988) Biol. Chem. Hoppe-Seyler 369, 247-250) comprising the C-terminal portion of the coding region and the 3'UTR, as well as a polymer chain reaction PCR fragment (Rapp et al., (1989) Biol. Chem. Hoppe-Seyler 370, 1071-1075) covering the missing part of the coding region from the amino-terminus.

Base Sequence↗
PubMed1991

Time-resolved cryo-electron microscopic study of the dissociation of actomyosin induced by photolysis of photolabile nucleotides.

The rapid release of a substrate or other ligand from photolabile precursors in a thin layer suspension of biological specimens followed by rapid freezing provides a method of trapping and visualizing short-lived states in a dynamic system. We demonstrate here the first successful application of this method to study the interaction of actin filaments with myosin subfragment 1 (S1) after release of nucleotides. The results obtained suggest that structural changes in actin filaments occur as a result of interaction with S1.

Actins↗