17 results on '"Brasseur R"'
Search Results
2. The topology of the S protein in the yeast-derived hepatitis B surface antigen particles.
- Author
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Sonveaux, N, primary, Conrath, K, additional, Capiau, C, additional, Brasseur, R, additional, Goormaghtigh, E, additional, and Ruysschaert, J M, additional
- Published
- 1994
- Full Text
- View/download PDF
3. Human apolipoprotein B signal sequence variants confer a secretion-defective phenotype when expressed in yeast.
- Author
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Sturley, S.L., primary, Talmud, P.J., additional, Brasseur, R., additional, Culbertson, M.R., additional, Humphries, S.E., additional, and Attie, A.D., additional
- Published
- 1994
- Full Text
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4. The 19-27 amino acid segment of gp51 adopts an amphiphilic structure and plays a key role in the fusion events induced by bovine leukemia virus.
- Author
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Vonèche, V, primary, Callebaut, I, additional, Kettmann, R, additional, Brasseur, R, additional, Burny, A, additional, and Portetelle, D, additional
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- 1992
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5. Differentiation of lipid-associating helices by use of three-dimensional molecular hydrophobicity potential calculations
- Author
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Brasseur, R., primary
- Published
- 1991
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6. The crystal structure of a penicilloyl-serine transferase of intermediate penicillin sensitivity. The DD-transpeptidase of streptomyces K15.
- Author
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Fonzé, E, Vermeire, M, Nguyen-Distèche, M, Brasseur, R, and Charlier, P
- Abstract
The serine DD-transpeptidase/penicillin-binding protein of Streptomyces K15 catalyzes peptide bond formation in a way that mimics the penicillin-sensitive peptide cross-linking reaction involved in bacterial cell wall peptidoglycan assembly. The Streptomyces K15 enzyme is peculiar in that it can be considered as an intermediate between classical penicillin-binding proteins, for which benzylpenicillin is a very efficient inactivator, and the resistant penicillin-binding proteins that have a low penicillin affinity. With its moderate penicillin sensitivity, the Streptomyces K15 DD-transpeptidase would be helpful in the understanding of the structure-activity relationship of this penicillin-recognizing protein superfamily. The structure of the Streptomyces K15 enzyme has been determined by x-ray crystallography at 2.0-A resolution and refined to an R-factor of 18.6%. The fold adopted by this 262-amino acid polypeptide generates a two-domain structure that is close to those of class A beta-lactamases. However, the Streptomyces K15 enzyme has two particular structural features. It lacks the amino-terminal alpha-helix found in the other penicilloyl-serine transferases, and it exhibits, at its surface, an additional four-stranded beta-sheet. These two characteristics might serve to anchor the enzyme in the plasma membrane. The overall topology of the catalytic pocket of the Streptomyces K15 enzyme is also comparable to that of the class A beta-lactamases, except that the Omega-loop, which bears the essential catalytic Glu(166) residue in the class A beta-lactamases, is entirely modified. This loop adopts a conformation similar to those found in the Streptomyces R61 DD-carboxypeptidase and class C beta-lactamases, with no equivalent acidic residue.
- Published
- 1999
7. Calculation of the three-dimensional structure of Saccharomyces cerevisiae cytochrome b inserted in a lipid matrix.
- Author
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Brasseur, R
- Abstract
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.
- Published
- 1988
- Full Text
- View/download PDF
8. Secondary Structure of Diphtheria Toxin and Its Fragments Interacting with acidic Liposomes Studied by Polarized Infrared Spectroscopy
- Author
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Cabiaux, V, Brasseur, R, Wattiez, R, Falmagne, P, Ruysschaert, J M, and Goormaghtigh, E
- Abstract
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-α-dipalmitoylphosphatidylcholine-DL-α-dipalmitoylphosphatidic acid liposomes at pH 7.3 results in a 10% increase in its α-helix content. At pH 4, in the presence of liposomes, the secondary structure of DT is characterized by the appearance of a β-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 α-helix content of CB1 increases by 10%, and polarization measurements indicate that the α-helices of CB1 at pH 4 are oriented parallel to the lipid acyl chains. On the other hand, the α-helix content of CB4 decreases dramatically while the low frequency β-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.
- Published
- 1989
- Full Text
- View/download PDF
9. Fusogenic properties of the C-terminal domain of the Alzheimer beta-amyloid peptide.
- Author
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Pillot, T, Goethals, M, Vanloo, B, Talussot, C, Brasseur, R, Vandekerckhove, J, Rosseneu, M, and Lins, L
- Abstract
A series of natural peptides and mutants, derived from the Alzheimer beta-amyloid peptide, was synthesized, and the potential of these peptides to induce fusion of unilamellar lipid vesicles was investigated. These peptide domains were identified by computer modeling and correspond to respectively the C-terminal (e.g. residues 29-40 and 29-42) and a central domain (13-28) of the beta-amyloid peptide. The C-terminal peptides are predicted to insert in an oblique way into a lipid membrane through their N-terminal end, while the mutants are either parallel or perpendicular to the lipid bilayer. Peptide-induced vesicle fusion was demonstrated by several techniques, including lipid-mixing and core-mixing assays using pyrene-labeled vesicles. The effect of peptide elongation toward the N-terminal end of the entire beta-amyloid peptide was also investigated. Peptides corresponding to residues 22-42 and 12-42 were tested using the same techniques. Both the 29-40 and 29-42 beta-amyloid peptides were able to induce fusion of unilamellar lipid vesicles and calcein leakage, and the amyloid 29-42 peptide was the most potent fusogenic peptide. Neither the two mutants or the 13-28 beta-amyloid peptide had any fusogenic activity. Circular dichroism measurements showed an increase of the alpha-helical content of the two C-terminal peptides at increasing concentrations of trifluoroethanol, which was accompanied by an increase of the fusogenic potential of the peptides. Our data suggest that the alpha-helical content and the angle of insertion of the peptide into a lipid bilayer are critical for the fusogenic activity of the C-terminal domain of the amyloid peptide. The differences observed between the fusogenic capacity of the amyloid 29-40 and 29-42 peptides might result from differences in the degree of penetration of the peptides into the membrane and the resulting membrane destabilization. The longer peptides, residues 22-42 and 12-42, had decreased, but significant, fusogenic properties associated with perturbation of the membrane permeability. These data suggest that the fusogenic properties of the C-terminal domain of the beta-amyloid peptide might contribute to the cytotoxicity of the peptide by destabilizing the cell membrane.
- Published
- 1996
10. Characterization of the interfacial behavior and structure of the signal sequence of Escherichia coli outer membrane pore protein PhoE.
- Author
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Batenburg, A M, Brasseur, R, Ruysschaert, J M, van Scharrenburg, G J, Slotboom, A J, Demel, R A, and de Kruijff, B
- Abstract
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.
- Published
- 1988
- Full Text
- View/download PDF
11. The role of proline in the membrane re-entrant helix of caveolin-1.
- Author
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Aoki S, Thomas A, Decaffmeyer M, Brasseur R, and Epand RM
- Subjects
- Amino Acid Substitution, Animals, Caveolin 1 genetics, Cell Line, Humans, Mice, Mutation, Missense, Proline genetics, Protein Structure, Secondary, Caveolae metabolism, Caveolin 1 metabolism, Endocytosis physiology, Proline metabolism
- Abstract
Caveolin-1 has a segment of hydrophobic amino acids comprising approximately residues 103-122. We have performed an in silico analysis of the conformational preference of this segment of caveolin-1 using PepLook. We find that there is one main group of stable conformations corresponding to a hydrophobic U bent model that would not traverse the membrane. Furthermore, the calculations predict that substituting the Pro(110) residue with an Ala will change the conformation to a straight hydrophobic helix that would traverse the membrane. We have expressed the P110A mutant of caveolin-1, with a FLAG tag at the N terminus, in HEK 293 cells. We evaluate the topology of the proteins with confocal immunofluorescence microscopy in these cells. We find that FLAG tag at the N terminus of the wild type caveolin-1 is not reactive with antibodies unless the cell membrane is permeabilized with detergent. This indicates that in these cells, the hydrophobic segment of this protein is not transmembrane but takes up a bent conformation, making the protein monotopic. In contrast, the FLAG tag at the N terminus of the P110A mutant is equally exposed to antibodies, before and after membrane permeabilization. We also find that the P110A mutation causes a large reduction of endocytosis of caveolae, cellular lipid accumulation, and lipid droplet formulation. In addition, we find that this mutation markedly reduces the ability of caveolin-1 to form structures with the characteristic morphology of caveolae or to partition into the detergent-resistant membranes of these cells. Thus, the single Pro residue in the membrane-inserting segment of caveolin-1 plays an important role in both the membrane topology and localization of the protein as well as its functions.
- Published
- 2010
- Full Text
- View/download PDF
12. Mode of membrane interaction and fusogenic properties of a de novo transmembrane model peptide depend on the length of the hydrophobic core.
- Author
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Lorin A, Charloteaux B, Fridmann-Sirkis Y, Thomas A, Shai Y, and Brasseur R
- Subjects
- Alanine chemistry, Amino Acids chemistry, Animals, Cattle, Cell Membrane metabolism, Hydrophobic and Hydrophilic Interactions, Leucine chemistry, Lipids chemistry, Liposomes metabolism, Models, Molecular, Protein Structure, Tertiary, Spectrophotometry, Infrared, Spectroscopy, Fourier Transform Infrared, Membranes chemistry, Peptides chemistry
- Abstract
Model peptides composed of alanine and leucine residues are often used to mimic single helical transmembrane domains. Many studies have been carried out to determine how they interact with membranes. However, few studies have investigated their lipid-destabilizing effect. We designed three peptides designated KALRs containing a hydrophobic stretch of 14, 18, or 22 alanines/leucines surrounded by charged amino acids. Molecular modeling simulations in an implicit membrane model as well as attenuated total reflection-Fourier transform infrared analyses show that KALR is a good model of a transmembrane helix. However, tryptophan fluorescence and attenuated total reflection-Fourier transform infrared spectroscopy indicate that the extent of binding and insertion into lipids increases with the length of the peptide hydrophobic core. Although binding can be directly correlated to peptide hydrophobicity, we show that insertion of peptides into a membrane is determined by the length of the peptide hydrophobic core. Functional studies were performed by measuring the ability of peptides to induce lipid mixing and leakage of liposomes. The data reveal that whereas KALR14 does not destabilize liposomal membranes, KALR18 and KALR22 induce 40 and 50% of lipid-mixing, and 65 and 80% of leakage, respectively. These results indicate that a transmembrane model peptide can induce liposome fusion in vitro if it is long enough. The reasons for the link between length and fusogenicity are discussed in relation to studies of transmembrane domains of viral fusion proteins. We propose that fusogenicity depends not only on peptide insertion but also on the ability of peptides to destabilize the two leaflets of the liposome membrane.
- Published
- 2007
- Full Text
- View/download PDF
13. Anti-hemostatic effects of a serpin from the saliva of the tick Ixodes ricinus.
- Author
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Prevot PP, Adam B, Boudjeltia KZ, Brossard M, Lins L, Cauchie P, Brasseur R, Vanhaeverbeek M, Vanhamme L, and Godfroid E
- Subjects
- Amino Acid Sequence, Animals, Humans, Insect Proteins chemistry, Insect Proteins genetics, Models, Biological, Models, Molecular, Molecular Sequence Data, Recombinant Proteins genetics, Recombinant Proteins metabolism, Sequence Alignment, Serpins chemistry, Serpins genetics, Hemostasis physiology, Insect Proteins metabolism, Ixodes chemistry, Saliva chemistry, Serpins metabolism
- Abstract
Serpins (serine protease inhibitors) are a large family of structurally related proteins found in a wide variety of organisms, including hematophagous arthropods. Protein analyses revealed that Iris, previously described as an immunomodulator secreted in the tick saliva, is related to the leukocyte elastase inhibitor and possesses serpin motifs, including the reactive center loop (RCL), which is involved in the interaction between serpins and serine proteases. Only serine proteases were inhibited by purified recombinant Iris (rIris), whereas mutants L339A and A332P were found devoid of any protease inhibitory activity. The highest Ka was observed with human leukocyte-elastase, suggesting that elastase-like proteases are the natural targets of Iris. In addition, mutation M340R completely changed both Iris substrate specificity and affinity. This likely identified Met-340 as amino acid P1 in the RCL. The effects of rIris and its mutants were also tested on primary hemostasis, blood clotting, and fibrinolysis. rIris increased platelet adhesion, the contact phase-activated pathway of coagulation, and fibrinolysis times in a dose-dependent manner, whereas rIris mutant L339A affected only platelet adhesion. Taken together, these results indicate that Iris disrupts coagulation and fibrinolysis via the anti-proteolytic RCL domain. One or more other domains could be responsible for primary hemostasis inhibition. To our knowledge, this is the first ectoparasite serpin that interferes with both hemostasis and the immune response.
- Published
- 2006
- Full Text
- View/download PDF
14. Role of the lid hydrophobicity pattern in pancreatic lipase activity.
- Author
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Thomas A, Allouche M, Basyn F, Brasseur R, and Kerfelec B
- Subjects
- Algorithms, Binding Sites, Catalysis, Dose-Response Relationship, Drug, Humans, Hydrogen-Ion Concentration, Hydrolysis, Kinetics, Lipase chemistry, Models, Molecular, Models, Statistical, Mutation, Olive Oil, Peptides chemistry, Plant Oils, Plasmids metabolism, Point Mutation, Protein Conformation, Protein Structure, Secondary, Triglycerides chemistry, Lipase metabolism, Pancreas enzymology
- Abstract
Pancreatic lipase is a soluble globular protein that must undergo structural modifications before it can hydrolyze oil droplets coated with bile salts. The binding of colipase and movement of the lipase lid open access to the active site. Mechanisms triggering lid mobility are unclear. The *KNILSQIVDIDGI* fragment of the lid of the human pancreatic lipase is predicted by molecular modeling to be a tilted peptide. Tilted peptides are hydrophobicity motifs involved in membrane fusion and more globally in perturbations of hydrophobic/hydrophilic interfaces. Analysis of this lid fragment predicts no clear consensus of secondary structure that suggests that its structure is not strongly sequence determined and could vary with environment. Point mutations were designed to modify the hydrophobicity profile of the [240-252] fragment and their consequences on the lipase-mediated catalysis were tested. Two mutants, in which the tilted peptide motif was lost, also have poor activity on bile salt-coated oil droplets and cannot be reactivated by colipase. Conversely, one mutant in which a different tilted peptide is created retains colipase dependence. These results suggest that the tilted hydrophobicity pattern of the [240-252] fragment is neither important for colipase binding to lipase, nor for interfacial binding but is important to trigger the maximal catalytic efficiency of lipase in the presence of bile salt.
- Published
- 2005
- Full Text
- View/download PDF
15. The human VPAC1 receptor: three-dimensional model and mutagenesis of the N-terminal domain.
- Author
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Lins L, Couvineau A, Rouyer-Fessard C, Nicole P, Maoret JJ, Benhamed M, Brasseur R, Thomas A, and Laburthe M
- Subjects
- Adenylyl Cyclases metabolism, Amino Acid Sequence, Animals, CHO Cells, Cricetinae, DNA, Complementary metabolism, Databases, Factual, Enzyme Activation, Epitopes, Humans, Kinetics, Ligands, Microscopy, Fluorescence, Models, Molecular, Molecular Sequence Data, Mutagenesis, Mutagenesis, Site-Directed, Mutation, Phenotype, Protein Binding, Protein Structure, Secondary, Protein Structure, Tertiary, Receptors, Vasoactive Intestinal Polypeptide, Type I, Sequence Homology, Amino Acid, Signal Transduction, Software, Structure-Activity Relationship, Transfection, Tryptophan metabolism, Receptors, Vasoactive Intestinal Peptide chemistry
- Abstract
The human VPAC(1) receptor for vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase activating peptide belongs to the class II family of G-protein-coupled receptors with seven transmembrane segments. Like for all class II receptors, the extracellular N-terminal domain of the human VPAC(1) receptor plays a predominant role in peptide ligand recognition. To determine the three-dimensional structure of this N-terminal domain (residues 1-144), the Protein Data Bank (PDB) was screened for a homologous protein. A subdomain of yeast lipase B was found to have 27% sequence identity and 50% sequence homology with the N-terminal domain (8) of the VPAC(1) receptor together with a good alignment of the hydrophobic clusters. A model of the N-terminal domain of VPAC(1) receptor was thus constructed by homology. It indicated the presence of a putative signal sequence in the N-terminal extremity. Moreover, residues (Glu(36), Trp(67), Asp(68), Trp(73), and Gly(109)) which were shown to be crucial for VIP binding are gathered around a groove that is essentially negatively charged. New putatively important residues for VIP binding were suggested from the model analysis. Site-directed mutagenesis and stable transfection of mutants in CHO cells indicated that Pro(74), Pro(87), Phe(90), and Trp(110) are indeed important for VIP binding and activation of adenylyl cyclase activation. Combination of molecular modeling and directed mutagenesis provided the first partial three-dimensional structure of a VIP-binding domain, constituted of an electronegative groove with an outspanning tryptophan shell at one end, in the N-terminal extracellular region of the human VPAC(1) receptor.
- Published
- 2001
- Full Text
- View/download PDF
16. Interaction between the N-terminal domain of gastric H,K-ATPase and the spectrin binding domain of ankyrin III.
- Author
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Festy F, Robert JC, Brasseur R, and Thomas A
- Subjects
- Amino Acid Sequence, Animals, Ankyrins metabolism, Binding Sites, Chromatography, Affinity, H(+)-K(+)-Exchanging ATPase genetics, H(+)-K(+)-Exchanging ATPase metabolism, Humans, Mice, Molecular Sequence Data, Precipitin Tests, Rabbits, Ankyrins chemistry, H(+)-K(+)-Exchanging ATPase chemistry, Spectrin metabolism, Stomach enzymology
- Abstract
We screened a cDNA bank of rabbit gastric fundic mucosa by two-hybrid assays looking for binding partners of the N-terminal domain of the rabbit gastric H,K-ATPase. We extracted five clones sharing more than 90% sequence identity. The longest clone codes for a protein sharing a high identity (96 and 96.8%, respectively) with a fragment of the membrane domain, from Arg-835 to Ser-873, plus the major part of the "spectrin binding domain" going from Glu-874 to Leu-1455 of human and mouse ankyrin III. We conclude that the membrane and spectrin binding domains of the rabbit ankyrin III are candidates for the binding partner of the N-terminal domain of the rabbit gastric H,K-ATPase. To validate the ankyrin-ATPase interaction and to test its specificity, we produced both domains in yeast and bacteria, coimmunoprecipitated them with an anti-ATPase antibody, and copurified them by affinity chromatography. The sequence of rabbit ankyrin III was not known, and this is the first report demonstrating that the ankyrin III and the H,K-ATPase interact with no intermediate. The interaction involves the N-terminal domain of the ATPase on one hand and the spectrin binding domain of the ankyrin on the other.
- Published
- 2001
- Full Text
- View/download PDF
17. Identification of key residues for interaction of vasoactive intestinal peptide with human VPAC1 and VPAC2 receptors and development of a highly selective VPAC1 receptor agonist. Alanine scanning and molecular modeling of the peptide.
- Author
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Nicole P, Lins L, Rouyer-Fessard C, Drouot C, Fulcrand P, Thomas A, Couvineau A, Martinez J, Brasseur R, and Laburthe M
- Subjects
- Adenylyl Cyclases metabolism, Amino Acid Sequence, Binding Sites, Computer Simulation, Humans, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, Protein Binding, Receptors, Vasoactive Intestinal Peptide genetics, Receptors, Vasoactive Intestinal Peptide, Type II, Receptors, Vasoactive Intestinal Polypeptide, Type I, Recombinant Proteins metabolism, Structure-Activity Relationship, Receptors, Vasoactive Intestinal Peptide agonists, Receptors, Vasoactive Intestinal Peptide metabolism, Vasoactive Intestinal Peptide analogs & derivatives, Vasoactive Intestinal Peptide metabolism
- Abstract
The widespread neuropeptide vasoactive intestinal peptide (VIP) has two receptors VPAC(1) and VPAC(2). Solid-phase syntheses of VIP analogs in which each amino acid has been changed to alanine (Ala scan) or glycine was achieved and each analog was tested for: (i) three-dimensional structure by ab initio molecular modeling; (ii) ability to inhibit (125)I-VIP binding (K(i)) and to stimulate adenylyl cyclase activity (EC(50)) in membranes from cell clones stably expressing human recombinant VPAC(1) or VPAC(2) receptor. The data show that substituting residues at 14 positions out of 28 in VIP resulted in a >10-fold increase of K(i) or EC(50) at the VPAC(1) receptor. Modeling of the three-dimensional structure of native VIP (central alpha-helice from Val(5) to Asn(24) with random coiled N and C terminus) and analogs shows that substitutions of His(1), Val(5), Arg(14), Lys(15), Lys(21), Leu(23), and Ile(26) decreased biological activity without altering the predicted structure, supporting that those residues directly interact with VPAC(1) receptor. The interaction of the analogs with human VPAC(2) receptor is similar to that observed with VPAC(1) receptor, with three remarkable exceptions: substitution of Thr(11) and Asn(28) by alanine increased K(i) for binding to VPAC(2) receptor; substitution of Tyr(22) by alanine increased EC(50) for stimulating adenylyl cyclase activity through interaction with the VPAC(2) receptor. By combining 3 mutations at positions 11, 22, and 28, we developed the [Ala(11,22,28)]VIP analog which constitutes the first highly selective (>1,000-fold) human VPAC(1) receptor agonist derived from VIP ever described.
- Published
- 2000
- Full Text
- View/download PDF
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