PubMed Health⌕ Search

Biomedical subjects

E Goormaghtigh

Publications and source records attributed to E Goormaghtigh.

At least 73 records · Page 4Linked to original sources
PubMed1991

Secondary structure and orientation of the amphipathic peptide GALA in lipid structures. An infrared-spectroscopic approach.

GALA, a synthetic, amphipathic 30-amino-acid peptide, based upon a Glu-Ala-Leu-Ala motive, was designed to mimic the behavior of viral fusion proteins. GALA is a water-soluble peptide with an aperiodic conformation at neutral pH, and becomes an amphipathic alpha helix as the pH is lowered to 5, where it interacts with phospholipid bilayers. Attenuated total-reflection infrared spectroscopy, using polarized light, provides information on the structure and orientation of the peptide and the lipids, which is not subject to artifacts due to light scattering with large particles. H/2H-exchange rate of the amide N-H group and analysis of the shape of the amide I' by Fourier self-deconvolution and curve fitting indicate that the alpha-helical content increases from 19% to 69%, on lowering the pH. A further increase to 100% alpha helix is observed after interaction with palmitoyloleoylglycerophosphocholine (PamOleGroPCho) vesicles. Dichroism data obtained with oriented bilayers of the PamOleGroPCho-GALA complex demonstrate that PamOleGroPCho hydrocarbon chains and the peptide alpha helical axis are essentially perpendicular (+/- 15) to the membrane plane. At neutral pH, in the presence of dimyristoylglycerophosphocholine (Myr2GroPCho), GALA is known to form discoidal structures similar to those formed under the same conditions by apolipoproteins AI and AII. In these discoidal complexes, the alpha-helical content was estimated to be 65%, with the rest of the structure being essentially unordered. No significant modification of the all-trans conformation of the hydrocarbon chain of Myr2GroPCho was detected upon disc formation. Dichroism measurements show that the alpha-helical axis is essentially parallel to the hydrocarbon chains. These data support a model in which a discoidal patch of the bilayer is surrounded by amphipathic helices which shield the hydrophobic region of the bilayer from the aqueous environment. The infrared spectrum of GALA in this complex was found to be very similar to those of apolipoproteins AI and AII which form discoidal complexes with Myr2GroPCho, but the spectrum is quite different from that of apolipoprotein B100 in low-density lipoproteins, which does not form discoidal complexes.

Amino Acid Sequence↗
PubMed1991

Spectroscopic investigation of structure in octarellin (a de novo protein designed to adopt the alpha/beta-barrel packing).

We present here a spectroscopic structural characterization of octarellin, a recently reported de novo protein modelled on alpha/beta-barrel proteins [K. Goraj, A. Renard and J.A. Martial (1990) Protein Engng, 3, 259-266]. Infrared and Raman spectra analyses of octarellin's secondary structure reveal the expected percentage of alpha-helices (30%) and a higher beta-sheet content (40%) than predicted from the design. When the Raman spectra obtained with octarellin and native triosephosphate isomerase (a natural alpha/beta-barrel) are compared, similar percentages of secondary structures are found. Thermal denaturation of octarellin monitored by CD confirms that its secondary structures are quite stable, whereas its native-like tertiary fold is not. Tyrosine residues, predicted to be partially hidden from solvent, are actually exposed as revealed by Raman and UV absorption spectra. We conclude that the attempted alpha/beta-barrel conformation in octarellin may be loosely packed. The criteria used to design octarellin are discussed and improvements suggested.

Circular Dichroism↗
PubMed1991

Lipid-itraconazole interaction in lipid model membranes.

Itraconazole, a lipophilic, fungal sterol-biosynthesis inhibitor, does not disturb membrane organization parameters measured by differential scanning calorimetry and infrared spectroscopy. Conformational analysis studies suggest that the molecular volume and the position of itraconazole in the lipid membrane is similar to that of dipalmitoyl phosphatidylcholine. The mean energy of interaction between itraconazole and the phospholipid is -60.6 kJ mol-1 whereas this energy in the pure lipid matrix is -54.3 kJ mol-1. The mean molecular area of itraconazole calculated by projecting the molecule on the lipid-water interface is equal to that occupied by the pure lipid (60 A2/molecule).

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1990

Secondary structure and dosage of soluble and membrane proteins by attenuated total reflection Fourier-transform infrared spectroscopy on hydrated films.

Attenuated total reflection Fourier-transform infrared spectroscopy of thin hydrated films of soluble and membrane protein included in a phospholipid bilayer is shown to provide useful information as to the secondary structure of the protein. The analysis of the amide I band of deuterated samples by Fourier self-deconvolution followed by a curve fitting was performed by a new procedure in which all the input parameters are generated by the computer rather than by the investigator. The results of this analysis provide a correct estimation of the alpha-helix and beta-sheet structure content with a standard deviation of 8.6% when X-ray structures are taken as a reference. We also show that the orientation of the different secondary structures resolved by the Fourier self-deconvolution/curve-fitting procedure and of the phospholipid acyl chains can be simultaneously evaluated for membrane proteins reconstituted in a lipid bilayer. Of special interest for reconstitution of membrane proteins, the lipid/protein ratio can be accurately and quickly determined from the infrared spectrum.

Electronic Data Processing↗
PubMed1990

Lipid and peptide specificities in signal peptide--lipid interactions in model membranes.

The present data show the critical importance of the anionic lipid content in monomolecular layers for the interaction with PhoE signal peptide. At 37 degrees C and 100 mM NaCl the interaction is maximal at 30-40 mol% anionic lipid. The results correlate with the reduced translocation competence of Escherichia coli strain HD3122, which has a much lower anionic lipid content as compared to the wild-type strain SD12 (De Vrije et al. (1988) Nature 334, 173-175). PhoE signal peptide analogs as N-formyl PhoE signal peptide, PhoE signal peptide +(1-7) and PhoE signal peptide Val-8----Trp-8 show the same lipid preference as PhoE signal peptide. On the other hand the affinity for an anionic lipid interface is strongly reduced for PhoE signal peptide Lys-19,-20----Asp-19,-20, which correlates with the less efficient translocation of PhoE protein carrying this signal sequence. At limiting anionic lipid concentrations there is a temperature and salt effect on the observed interaction, which is related to a conformational change of the peptide. Signal sequences show clearly conformational flexibility in responds to environmental conditions. Under the conditions used in this study FTIR spectra of PhoE signal peptide-DOPG monolayers show a high content of beta-structure and beta-turn.

Amino Acid Sequence↗
PubMed1990

Mode of assembly of amphipathic helical segments in model high-density lipoproteins.

The structure of discoidal apo A-I-phospholipid complexes, representing the metabolic precursors of mature high-density lipoprotein particles, was studied by a combination of both a theoretical and an experimental approach. The secondary structure of the complex was determined by circular dichroic measurements, while the relative orientation of the apo A-I helical segments and of the phospholipid acyl chains was determined by ATR infrared measurements. Fluorescence energy transfer between the tryptophan residues of apo A-I and fluorescent phospholipid probes yielded an estimation of the relative topography of the lipid and apolipoprotein components in discoidal and spherical particles. The theoretical approach consisted of the identification of the helical segments in various apo A-I species. These segments were then oriented at a lipid/water interface by minimization of their hydrophobic and hydrophilic transfer energies. The calculation of the hydrophobicity profiles along the axis of the helices leads to the identification of specific interactions between pairs of helices. The helices were further assembled together with the phospholipids by computer modelling, enabling an estimation of the dimensions of the complex. The combination of the experimental and theoretical results yielded a model for discoidal apolipoprotein-phospholipid complexes, in which the amphipathic helical segments are oriented along the edges of the discs. Such a model can be extended to the conversion of these complexes into mature spherical HDL, through the formation of a cholesteryl ester core.

Amino Acid Sequence↗
PubMed1990

Structure of the adriamycin-cardiolipin complex. Role in mitochondrial toxicity.

Adriamycin and its derivatives are among the most efficient antimitotics used in clinical therapy. A specific cardiotoxicity places a limit on the total dose of adriamycin that may be administered. The mechanism of cardiac toxicity is complex. Data accumulated from in vitro and in vivo studies indicate a possible common cause for the inhibition of numerous enzymes and tissue degradation by a free radical mechanism: the binding of adriamycin to the inner mitochondrial membrane cardiolipin. The structure of the adriamycin-cardiolipin complex has been investigated by using physico-chemical techniques and via conformational analysis. The results open a rational way to design new structures that are less cardiotoxic.

Animals↗
PubMed1989

Evaluation of the secondary structure of apo B-100 in low-density lipoprotein (LDL) by infrared spectroscopy.

The secondary structure of the apo B-100 protein present in human low density lipoprotein has been investigated by transmission and attenuated total reflection infrared spectroscopy. The amount of beta-sheet (41%) is significantly higher than that determined by CD spectroscopy in the present study (12%) and elsewhere (15-16%). The high percentage of beta-sheet structure in apo B-100 supports the importance of such segments in maintaining the lipid-protein assembly in LDL. Polarized infrared spectroscopy indicates that the beta-sheet component of apo B-100 adopts a preferential orientation with respect to the phospholipid monolayer surrounding the LDL, whereas no such orientation is observed for the other secondary structure components.

Apolipoprotein B-100↗
PubMed1989

Secondary structure of diphtheria toxin and its fragments interacting with acidic liposomes studied by polarized infrared spectroscopy.

We used infrared attenuated total reflection spectroscopy to study the structure of diphtheria toxin (DT) and its fragments A, B, CB1, and CB4 as a function of the pH in the absence and in the presence of phospholipid vesicles. Binding of DT to asolectin or DL-alpha-dipalmitoylphosphatidylcholine-DL-alpha-dipalmitoylphosphatidic acid liposomes at pH 7.3 results in a 10% increase in its alpha-helix content. At pH 4, in the presence of liposomes, the secondary structure of DT is characterized by the appearance of a beta-sheet structure with strengthened hydrogen bonds which did not exist before pH lowering. DT fragment B displays little conformational change upon pH lowering in the presence of liposomes. However, the alpha-helix content of CB1 increases by 10%, and polarization measurements indicate that the alpha-helices of CB1 at pH 4 are oriented parallel to the lipid acyl chains. On the other hand, the alpha-helix content of CB4 decreases dramatically while the low frequency beta-sheet content increases. Dichroism measurements demonstrate that this sheet lies close to a parallel to the bilayer surface. The fragment A of DT experiences a large conformational change upon pH lowering and binds to the liposome membrane even in the absence of DT fragment B. The conformational modification of DT fragment A is fully reversed when pH is brought back to 7.3.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1989

Secondary structure and orientation of a chemically synthesized mitochondrial signal sequence in phospholipid bilayers.

A pre-sequence of 25 amino acids is required for import of yeast cytochrome oxidase subunit IV into mitochondria. Structure and orientation of the 25 amino acids synthesized peptide (p25) in a lipid bilayer were investigated by infrared attenuated total reflection spectroscopy. This method allowed to overcome the difficulties related to the optical turbidity due to the light scattering on membrane fragments which prevents the use of circular dichroism. We demonstrate here that incubation of the peptide with DOPC (dioleoylphosphatidylcholine) and DOPC-CL (dioleoylphosphatidylcholine - cardiolipin) liposomes is accompanied by an increase in alpha-helical content as compared to beta structure. Polarisation measurements indicate that the amphipathic helical segment is inserted parallel to the lipid acyl chains in cardiolipin containing liposomes.

Cardiolipins↗
PubMed1989

Secondary structure changes of diphtheria toxin interacting with asolectin liposomes: an infrared spectroscopy study.

Until now, the study of the secondary structure of diphtheria toxin (DT) in the presence of phospholipid vesicles as a function of the pH has been prevented by the optical turbidity of the solution. In the present paper, this problem has been overcome by the use of IR attenuated total reflection (IR-ATR) spectroscopy. Incubation of DT with asolectin liposomes at pH 7.3 results in the binding of DT on to the liposomes and in a 10% increase in its alpha-helix content. At pH 4, in the presence of asolectin liposomes, the secondary structure of DT is characterized by the appearance of a beta-sheet structure with strengthened hydrogen bonds, which did not exist before lowering of the pH. This new type of beta-sheet (low frequency beta-sheet) involves 25% of the amino acid residues of the protein.

Binding, Competitive↗
PubMed1988

Mode of insertion of praziquantel and derivatives into lipid membranes.

We report here about the relationship between the destabilization of the lipid organization induced by praziquantel and derivatives and their mode of insertion into the lipid matrix. Measurements of lipid transition temperature and efflux of 6-carboxyfluorescein encapsulated in liposomes establish the lipid destabilizing capacity of praziquantel as compared to praziquantel derivatives. IR spectroscopy (attenuated total reflection technique) applied to oriented lipid bilayers indicates that praziquantel or derivatives do not modify significantly the lipid structure. In order to give a molecular description of the position of the drug into the lipid bilayer, we applied a conformational analysis procedure making it possible to calculate the structure of amphiphilic molecules assembled in aggregates and the mode of insertion of amphiphilic drugs into a lipid layer. The praziquantel lipid destabilizing capacity is explained in terms of the high praziquantel-lipid interaction and the large area occupied per drug molecule in the lipid layer.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1988

The mode of insertion of the paramyxovirus F1 N-terminus into lipid matrix, an initial step in host cell/virus fusion.

A protein sequence of the viral envelope is responsible for the fusion of the viral membrane with the cell membrane. It has been hypothesized that this fusion protein acts by inserting its hydrophobic moiety into the target membrane. The paper reports on the mode of insertion of the Newcastle disease virus (NDV) fusion protein N-terminus peptide into a lipid membrane using infrared spectroscopy and conformational analysis. The peptide was shown to adopt an oblique orientation with respect to the lipid acyl chains. This unusual orientation is expected to induce the disorganization of the surrounding lipids, which had been suggested as one of the initial events in the virus/cell fusion.

Amino Acid Sequence↗
PubMed1987

Study of the adriamycin-cardiolipin complex structure using attenuated total reflection infrared spectroscopy.

Adriamycin plays a prominent role in the treatment of leukemia and solid tumors in man. The mode of interaction of adriamycin with its nuclear target, responsible for its therapeutic effect, is known [Berman, H. M., & Young, P.R. (1981) Annu. Rev. Biophys. Bioeng. 10, 87-114]. The planar anthracycline moiety of adriamycin intercalates between the base pairs whereas the sugar moiety fits into the DNA large groove. However, the cardiotoxicity of adriamycin places a limit on the total dose that may be given [Minow, R. A., Banjamin, R.S., & Gottlieb, J. A. (1975) Cancer Chemother. Rep. 6, 195-202]. Much evidence suggests that the mitochondrial membrane could be the target responsible for adriamycin cardiotoxicity. The formation of a very stable complex between adriamycin and cardiolipin, a phospholipid specific to the inner mitochondrial membrane, has been shown to inhibit several mitochondrial membrane enzymes whose activities depend on the presence of cardiolipin. Using attenuated total reflection infrared spectroscopy, we demonstrate here that, in the adriamycin-cardiolipin complex, both cardiolipin and adriamycin structures are modified as compared with the pure substances. Dichroism values indicate a slight reorientation of the cardiolipin molecule toward a normal to the plane of the bilayer whereas adriamycin, which shows no ordering in a pure phase, is highly ordered in the complex, the anthracycline moiety titled at about 40 degrees with respect to the normal to the plane of the bilayer. The partial disappearance of NH3+ characteristic bands indicates the involvement of the positively charged amino group of adriamycin in the complex formation.

Ammonia↗
PubMed1987

A hexameric form of the Neurospora crassa plasma membrane H+-ATPase.

As isolated by our recently developed large-scale procedure, the Neurospora plasma membrane H+-ATPase exists as a homogeneous, oligomeric complex of 105,000-Da monomers with a molecular mass equivalent to a spherical protein of about 1 million Da, as judged by its behavior during chromatography on calibrated columns of Sepharose CL-6B and CL-4B. Treatment of this complex with the nonionic detergent, Tween 20, followed by Sepharose column chromatography in the presence of this detergent produces particles with an apparent molecular mass reduced by 100-300 kDa, and, importantly, when the isolated complex is treated with Tween 20 and then subjected to Sepharose chromatography in the absence of detergent, fully viable, largely detergent-free, homogeneous particles with a molecular mass equivalent to a spherical protein of 670,000 Da are formed. As assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, treatment of the particles isolated in the presence of Tween 20 with glutaraldehyde progressively yields dimers, trimers, tetramers, pentamers, and hexamers of the 105,000-Da monomer, with the expected precursor-product relationships, but no species larger than a hexamer is formed. These results thus strongly indicate that these particles are hexamers of 105,000-Da monomers. Glutaraldehyde crosslinking experiments with the ca. 1 million- and 670,000-Da particles indicate that they too are hexamers, suggesting that the differences in the apparent sizes of the three types of particles are most likely due to bound detergents. Possible implications of these findings are discussed.

Biological Transport, Active↗
PubMed1986

Density-based separation of liposomes by glycerol gradient centrifugation.

Sonicated liposomes of soybean phospholipids (asolectin) distribute nearly throughout a 19-22% (v/v) glycerol gradient when centrifuged to near equilibrium. Upon recentrifugation on an identical gradient, liposomes selected from several positions in such a gradient migrate as narrow bands to positions close to their original positions, indicating that the liposome distribution in the first gradient is the result of a density-based fractionation. Molecular sieve chromatography, turbidity, and trapped volume measurements indicate that the liposome densities are qualitatively related to their size, with the larger liposomes more dense than the smaller ones. Size estimates obtained by electron microscopy of negatively stained preparations indicate that the fractionation is effective for liposomes with diameters ranging from 200 to 600 A, with maximum efficiency in the range 200-300 A where the majority of the liposomes is found. Interestingly, high concentrations of liposomes improve the efficiency of the fractionation procedure. The size dependence of liposome density is shown not to be due to differential glycerol permeability or lipid composition, and is therefore most likely due to variations in the specific volumes of the individual phospholipid molecules owing to the curvature of the liposomes. Finally, freezing of the glycerol gradient fractions in liquid N2 and storage at -70 degrees C does not modify the size of the isolated liposomes. It is suggested that glycerol density gradient fractionation of liposomes could be a useful general method for obtaining liposomes of reasonably uniform size in large quantities and high concentrations.

Centrifugation, Density Gradient↗
PubMed1986

Mechanism of inhibition of mitochondrial enzymatic complex I-III by adriamycin derivatives.

We demonstrate here that complex I-III of bovine heart mitochondrial membrane is inhibited by adriamycin derivatives. This inhibition is a cardiolipin-dependent process. This lipid, specific to the inner mitochondrial membrane, has been shown previously to interact specifically with adriamycin in model membranes (Goormaghtigh, E., Chatelain, P., Caspers, J. and Ruysschaert, J.-M. (1980) Biochim. Biophys. Acta 597, 1-14) and in mitochondrial membranes (Cheneval, D., Müller, M., Toni, R., Ruetz, S. and Carafoli, E. (1985) J. Biol. Chem. 260, 13003-13007). The differential scanning calorimetry data indicate that, in multilamellar liposomes, the formation of antibiotic-cardiolipin complexes induces a clustering of cardiolipin molecules. Conformational analysis of the antibiotic-cardiolipin complexes suggests that plane-plane interactions between the antibiotics aromatic moieties stabilize this complex formation. Possible mechanisms of inactivation of complex I-III by adriamycin are proposed.

Animals↗