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Biomedical subjects

R Brasseur

Publications and source records attributed to R Brasseur.

At least 145 records · Page 8Linked to original sources
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

Antigenic variants of bovine leukemia virus (BLV) are defined by amino acid substitutions in the NH2 part of the envelope glycoprotein gp51.

Previous studies with monoclonal antibodies of the antigenic structure of bovine leukemia virus (BLV) envelope glycoprotein (gp51) have identified three epitopes (F, G, H) directly involved in the infectivity of BLV, F, G, and H lost their reactivity with the respective monoclonal antibodies after treatment with a reducing agent, indicating that these epitopes were conformational. Sequence comparisons between BLV mutants and differential reactivities of urokinase or proteinase K gp51 fragments with monoclonal antibodies indicated that the NH2 moiety of the env protein harbored the three architectural determinants F, G, and H. ELISA tests demonstrated that anti-F, -G, and -H monoclonal antibodies were maximally reactive toward intact virions whereas they showed much poorer affinities for their respective epitopes when presented on a purified protein. Accordingly, an efficient vaccine against BLV infection will include at least the identified gp51 region presented in its native architectural configuration.

Amino Acid Sequence↗
PubMed1989

A multicentre open trial of fluoxetine in depressed out-patients in Belgium.

Fluoxetine hydrochloride (ProzacR, Lilly), a new antidepressant that specifically inhibits serotonin reuptake was evaluated in an open comparative trial performed in 8 Belgian centres over a period of 9 months. Seventy-six outpatients were enrolled of which 66 were evaluable. Thirty-six patients received an 8 week fluoxetine treatment and 31 patients received tricyclic antidepressants (TCA). Patients from both treatment groups were evaluated for safety and the fluoxetine treated group of patients was assessed for efficacy. On the Hamilton Depression Scale, the mean score dropped from 26.4 to 7.0 after 8 weeks (p less than 0.0001). The Clinical and Patient's Global Impression scores showed a similar evolution. Patients treated with fluoxetine tended to report fewer side-effects than the TCA treated group. Therefore fluoxetine can be regarded as an efficient antidepressant which is at least as well tolerated as TCAs.

Antidepressive Agents, Tricyclic↗
PubMed1988

Calculation of the three-dimensional structure of Saccharomyces cerevisiae cytochrome b inserted in a lipid matrix.

Cytochrome b is an integral membrane protein, which forms the core of the ubiquinol-cytochrome c oxidoreductase (cytochrome bc1) complex. A computer-aided three-dimensional modeling procedure was carried out in four steps. First, the candidate hydrophobic helices were searched for throughout the protein primary sequence by a computer procedure based upon the method of Eisenberg; second, a secondary helical structure was imposed to the transmembrane peptides; third, the helical segments at a lipid-water interface were oriented, and finally the possible interactions between helices with similar properties were investigated. This procedure enabled the identification of nine hydrophobic segments, of which eight are membrane-spanning helices while one has amphipathic properties. Three hydrophilic receptor-binding domains were also identified. Based upon their hydrophobicity profiles, the transmembrane helices could be associated in pairs inside the lipid bilayer. In our folding model proposed for cytochrome b, all mutation sites are not only located on the same side of the membrane but are also in close proximity in the three-dimensional structure. Inhibitor resistance mutational sites which were recently characterized (di Rago, J.-P., and Colson, A.-M. (1988) J. Biol. Chem. 263, 12564-12570) have been located on this model. Moreover, the receptor-binding domains and the mutation sites are close neighbors in the three-dimensional spatial representation.

Amino Acid Sequence↗
PubMed1988

Conformational analysis of lipid-associating proteins in a lipid environment.

Two major types of helical structures have been identified in lipid-associating proteins, being either amphipathic or transmembrane domains. A conformational analysis was carried out to characterize some of the properties of these helices. These calculations were performed both on isolated helices and in a lipid environment. According to the results of this analysis, the orientation of the line joining the hydrophobic and hydrophilic centers of the helix seems to determine the orientation of the helix at the lipid/water interface. The calculation of this parameter should be useful to discriminate between an amphipathic helix, parallel to the interface and a transmembrane helix orientated perpendicularly. The membrane-spanning helices are completely immersed in the phospholipid bilayer and their length corresponds to about the thickness of the hydrophobic core of the DPPC bilayer. The energy of interaction, expressed per phospholipid is significantly higher for the transmembrane compared to the amphipathic helices. For the membrane-spanning helices the mean energy of interaction is higher than the interaction energy between two phospholipids, while it is lower for most amphipathic helices. This might account for the stability of these protein-anchoring domains. This computer modeling approach should usefully complement the statistical analysis carried out on these helices, based on their hydrophobicity and hydrophobic moment. It represents a more refined analysis of the domains identified by the prediction techniques and stress the functional character of lipid-associating domains in membrane proteins as well as in soluble plasma lipoproteins.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1988

A comparative model membrane study on structural effects of membrane-active positively charged anti-tumor drugs.

The interaction of a number of positively charged anti-tumor drugs with cardiolipin-containing model membranes has been investigated using 31P nuclear magnetic resonance (31P-NMR), differential scanning calorimetry (DSC) and monolayer techniques. It appeared that the ellipticines used (i.e., celiptium and 2-N-methylellipticinium), and also ethidium bromide, completely blocked Ca2+-induced HII phase formation in pure cardiolipin liposomes at molar ratios of drug-to-lipid of approx. 1:1. For the anthracyclines adriamycin and 4'-epi-adriamycin, a similar effect was observed, but now a 2:1 ratio was required. 31P-NMR experiments on dioleoylphosphatidylethanolamine/cardiolipin mixed liposomes indicated that the two anthracyclines, but not the other three drugs, were capable of inducing macroscopic phase separation into domains enriched in drug-cardiolipin complexes and domains enriched in the zwitterionic phospholipid species. DSC experiments on dipalmitoylphosphatidylcholine/cardiolipin mixtures led, with the exception of 2-N-methylellipticinium, to the same conclusion. Measurements of surface pressure and surface potential of cardiolipin monolayers at the air/water interface as well as conformational analysis of the various drug-cardiolipin recombinants showed that the ellipticines are deeply embedded in the acyl chain region of the bilayer, while the anthracyclines and ethidium bromide are preferentially localized in the interface. All drugs share an important electrostatic interaction with the negatively charged phosphates of cardiolipin.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1988

Conformational analysis of lipoxin A, lipoxin B and their trans-isomers.

Lipoxin A and lipoxin B (LXA and LXB) are formed from the oxygenation of arachidonic acid by interactions between the 5- and 15-lipoxygenases of human leukocytes. Each compound displays highly stereospecific biological actions. Here, we present a computational description of the following compounds: lipoxin A, (5S,6R,15S)-trihydroxy-7,9,13-trans-11-cis-eicosatetraenoic acid; 11-trans-lipoxin A, (5S,6R,15S)-trihydroxy-7,9,11,13-trans-eicosatetraenoic acid; lipoxin B, (5S,14R,15S)-trihydroxy-6,10,12-trans-8-cis-eicosatetraenoic acid; and 8-trans-lipoxin B, (5S,14R,15S)-trihydroxy-6,8,10,12-trans-eicosatetraenoic acid. The analyses considered van der Waals energy, electrostatic interactions, torsional potential, and alterations in electrostatic forces. Additional analyses were carried out with each of the four compounds forming complexes with one calcium ion. Each compound gave very different conformers. Both lipoxin A and lipoxin B can form globular conformations, while their all-trans isomers form rigid extended structures. When complexes with each of these compounds and one calcium ion were examined (i.e., (LXA)2Ca: (11-trans-LXA)2Ca), both LXA and LXB formed several flexible conformations including crumpled, wrapped or extended conformations. In this situation, LXA showed a higher probability than LXB to wrap around one Ca2+. In contrast, the two all-trans isomers always lead to extended conformations. Results from the present study illustrate that changes in the stereochemistry of LXA and LXB lead to unique conformations which may underlie the different biological actions of these compounds. Moreover, they indicate that the conformations of eicosanoids can change while in aqueous or hydrophobic environments (i.e., biomembranes).

Arachidonate 15-Lipoxygenase↗
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

Characterization of the interfacial behavior and structure of the signal sequence of Escherichia coli outer membrane pore protein PhoE.

The behavior of the chemically synthesized PhoE signal peptide and signal peptide fragments on hydrophilic-hydrophobic interfaces was studied with circular dichroism and monolayer techniques. The experimental results were compared with computer-calculated predictions of peptide structure, orientation, and molecular area. The complete signal sequence was found to aggregate in a beta-sheet structure when introduced in an aqueous environment; on the other hand, in sodium dodecyl sulfate micelles approximately 75% alpha-helical structure was observed. Assuming this to reflect the actual structure in a peptide monolayer and taking into account the orientations predicted for the fragments, the measured molecular areas suggest a looped orientation of the signal sequence with both N and C terminus in the water phase.

Bacterial Outer Membrane Proteins↗
PubMed1988

Computed conformational analysis of lipoxins and their ionic complexes.

The possible molecular conformations of four structurally and biologically different lipoxins derivatives were predicted by a systematic structure tree theoretical analysis. This method takes into account the London-Van der Waals energy of interaction, the electrostatic interaction, the rotation energy of the torsional angles and the energy of transfer through a possible lipid-water interface. Finally, the conformers derived from the structure tree and with a high probability of existence were submitted to the energy minimization procedure. The most probable conformers of lipoxin A: 5S,6R,15S-trihydroxy-7,9,13 trans-11 cis-eicosatetraenoic acid (LXA); 11 trans lipoxin A: 5S,6R,15S-trihydroxy-7,9,11,13 trans-eicosatetraenoic acid (11t-LXA); lipoxin B: 5S,14R 15S-trihydroxy-6,10,12 trans-8 cis-eicosatetraenoic acid (LXB) and 8 trans lipoxin B: 5S,14R,15S-trihydroxy- 6,8,10,12 trans-eicosatetraenoic acid (8t-LXB) in their isolated form or when forming complexes with one calcium ion are presented. The four isolated compounds lead to vastly different conformations. Lipoxin A can form the most globular conformer while lipoxin B seems to be slightly more extended. The all trans isomer of lipoxin B forms an extended conformer and 11 trans lipoxin A gives a fully extended molecule. Complexes of a pair of these compounds with one calcium ion were shown to lead to vastly different conformations. Both (LXA) 2Ca and (LXB)2Ca form crumpled or extended structure, the LXA molecules being more wrapped around Ca2+ than LXB molecules. The (11t-LXA)2Ca and (8t-LXB)2Ca complexes present a high probability of extended conformations. Our description merely shows that the peculiar stereochemistry of these molecules lead to equilibria between conformers or to very static conformers, the flexibility and rigidity of which being probably relevant in view of their different biological activities.

Chemical Phenomena↗
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↗
PubMed1988

Fusion (F) protein gene of Newcastle disease virus: sequence and hydrophobicity comparative analysis between virulent and avirulent strains.

The nucleotide and predicted amino acid sequences have been obtained for the fusion (F) protein gene of the avirulent strain La Sota of Newcastle disease virus (NDV). The F1 N-terminus begins with the tripeptide Leu-Ileu-Gly instead of Phe-X-Gly as usually observed in fusion peptide. It was found that the cleavage-activation domain of the avirulent La Sota strain contained single (but no pairs of) basic residues in the sequence Gly-Arg-Gln-Gly-Arg. Hydrophobicity analysis suggested that the cleavage-activation domain became more hydrophobic and could be less accessible for host-specific protease(s); dibasic residues next to the F1 N-terminus were shown to be important for keeping the cleavage-activation site in exposed positioning, suitable for F protein activation. Comparative sequence analysis of the NDV F proteins revealed a striking homology between lentogenic La Sota and mesogenic Beaudette C strains. Furthermore, 58 variable positions were recorded in the NDV F protein, excluding signal sequence; some of these mutations, in the cysteine-clustered region, were surmised to alter virulence.

Amino Acid Sequence↗
PubMed1988

Mode of insertion into a lipid membrane of the N-terminal HIV gp41 peptide segment.

The complete amino-acid sequence of the gp160 polyprotein of HIV (strain WMJ1) has been analyzed by the Eisenberg procedure. The region surrounding the cleavage site between the gp120 and the gp41 subunit contains a receptor-like region immediately followed by a transmembrane-like region containing approximately 13 residues. These two regions are separated by the cleavage site between gp120 and gp41. Since the same arrangement exists in some paramyxoviruses (unpublished observation) and since the effective cleavage between a receptor-like region and the transmembrane-like region is required in paramyxoviruses to generate fusogenic segment (located at the N-terminal sequence of the transmembrane-like region), we have focused our analysis on the conformational properties of the N-terminal peptide segment of HIV gp41. This peptide segment, which consists of a helical structure according to Garnier prediction, was oriented at the lipid-water interface using a theoretical analysis method that we recently developed. Analysis of the transmembrane peptide determined by Eisenberg method shows that the helical segment orients itself in the lipid monolayer obliquely with respect to the lipid-water interface. Since this rather unusual orientation for a membrane segment of a protein is also found in the fusogenic peptide of the Newcastle Disease Virus (Virus Genes, in press) and seems to possess membrane destabilizing properties, it is in agreement with previous reports suggesting a fusogenic role for the N-terminal part of gp41.

HIV↗
PubMed1987

Conformational analysis of gramicidin-gramicidin interactions at the air/water interface suggests that gramicidin aggregates into tube-like structures similar as found in the gramicidin-induced hexagonal HII phase.

The energetics of interaction and the type of aggregate structure in lateral assemblies of up to five gramicidin molecules in the beta 6.3 helical conformation at the air/water interface was calculated using conformational analysis procedures. It was found that within the aggregate two types of gramicidin interaction occur. One leading to a linear organization with a mean interaction energy between monomers of -6 kcal/mol and one in a perpendicular direction leading to a circularly organization with a lower mean interaction energy of -10 kcal/mol. Extrapolation towards larger gramicidin assemblies predicts that gramicidin itself could form tubular structures similar to those found in the gramicidin-induced HII phase. The tryptophans appear to play an essential role in the tubular organization of the gramicidin aggregate, since they determine the cone shape of the monomer and contribute to the structure of the monomer and oligomer by stacking interactions. These results, which are discussed in the light of experimental observations of gramicidin self-association in model membranes and the importance of the tryptophans for HII phase formation, further support the view (Killian, J.A. and De Kruijff, B. (1986) Chem. Phys. Lipids 40, 259-284) that gramicidin is a first example of a new class of hydrophobic polypeptides which can form cylindrical structures within the hydrophobic core of the membrane.

Gramicidin↗
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

Functional differentiation of amphiphilic helices of the apolipoproteins by hydrophobic moment analysis.

The amphiphilic character of different plasma apolipoproteins was investigated by a combination of established hydrophobicity analysis methods. These methods proved to be powerful in the detection of amphiphilic phospholipid-binding domains. Within this class of lipid-binding domains, lecithin-cholesterol acyltransferase activating and non-activating helices could be differentiated by calculating hydrophobic moments at different angles. We conclude that the hydrophobic characteristics of the different helices determined the mode of lipid binding and the substrate properties of these phospholipid-protein complexes for the lecithin-cholesterol acyltransferase reaction.

Apolipoprotein A-II↗