Benchmarking the PEPOP methods for mimicking discontinuous epitopes.

Background: Computational methods provide approaches to identify epitopes in protein Ags to help characterizing potential biomarkers identified by high-throughput genomic or proteomic experiments. PEPOP version 1.0 was developed as an antigenic or immunogenic peptide prediction tool.

Use of Phage Display technology in development of canine visceral leishmaniasis vaccine using synthetic peptide trapped in sphingomyelin/cholesterol liposomes.

Background: Leishmania parasites can cause visceral or cutaneous disease and are found in subtropical and tropical regions of the Old and New World. The pathology of the infection is determined by both host immune factors and species/strain differences of the parasite. Dogs represent the major reservoir of Leishmania infantum (syn.

Immunodiagnosis of canine visceral leishmaniasis using mimotope peptides selected from phage displayed combinatorial libraries.

ELISA and RIFI are currently used for serodiagnosis of canine visceral leishmaniasis (CVL). The accuracy of these tests is controversial in endemic areas where canine infections by Trypanosoma cruzi may occur. We evaluated the usefulness of synthetic peptides that were selected through phage display technique in the serodiagnosis of CVL.

Use of a synthetic biosensor for neutralizing activity-biased selection of monoclonal antibodies against atroxlysin-I, an hemorrhagic metalloproteinase from Bothrops atrox snake venom.

Background: The snake Bothrops atrox is responsible for the majority of envenomings in the northern region of South America. Severe local effects, including hemorrhage, which are mainly caused by snake venom metalloproteinases (SVMPs), are not fully neutralized by conventional serum therapy. Little is known about the immunochemistry of the P-I SVMPs since few monoclonal antibodies (mAbs) against these molecules have been obtained.

Identification of new sphingomyelinases D in pathogenic fungi and other pathogenic organisms.

Sphingomyelinases D (SMases D) or dermonecrotic toxins are well characterized in Loxosceles spider venoms and have been described in some strains of pathogenic microorganisms, such as Corynebacterium sp. After spider bites, the SMase D molecules cause skin necrosis and occasional severe systemic manifestations, such as acute renal failure. In this paper, we identified new SMase D amino acid sequences from various organisms belonging to 24 distinct genera, of which, 19 are new.

Discontinuous epitopes on the C2 domain of coagulation Factor VIII mapped by computer-designed synthetic peptides.

The occurrence of alloantibodies against Factor VIII (FVIII) is the main iatrogenic complication in haemophilia A (HA). Anti-FVIII autoantibodies may also spontaneously appear in non-HA patients, leading to acquired haemophilia A. In both contexts, the antibody response against FVIII is complex and difficult to analyse due to the lack of suitable tools.

Epitope spreading of the anti-CYP2D6 antibody response in patients with autoimmune hepatitis and in the CYP2D6 mouse model.

Autoimmune hepatitis (AIH) is a serious chronic inflammatory disease of the liver with yet unknown etiology and largely uncertain immunopathology. The hallmark of type 2 AIH is the generation of liver kidney microsomal-1 (LKM-1) autoantibodies, which predominantly react to cytochrome P450 2D6 (CYP2D6). The identification of disease initiating factors has been hampered in the past, since antibody epitope mapping was mostly performed using serum samples collected late during disease resulting in the identification of immunodominant epitopes not necessarily representing those involved in disease initiation.

Analysis of the variability of human normal urine by 2D-GE reveals a "public" and a "private" proteome.

The characterization of the normal urinary proteome is steadily progressing and represents a major interest in the assessment of clinical urinary biomarkers. To estimate quantitatively the variability of the normal urinary proteome, urines of 20 healthy people were collected. We first evaluated the impact of the sample conservation temperature on urine proteome integrity.

Association of genetic variations in TCF7L2, SLC30A8, HHEX, LOC387761, and EXT2 with Type 2 diabetes mellitus in Tunisia.

Aim: In recent genome-wide association studies, genetic variants in TCF7L2, SLC30A8, HHEX, LOC387761, and EXT2 were associated with risk for type 2 diabetes mellitus (T2DM). We aimed at investigating the association of these single-nucleotide polymorphisms (SNPs) with T2DM and defining their corresponding allelic and genotypic combinations in the Tunisian population. We also tried to determine the effect of R325W of SLC30A8 on the modeled structural properties of the protein.

Cardiotoxic effects of Loxosceles intermedia spider venom and the recombinant venom toxin rLiD1.

Loxosceles spider bites cause many human injuries worldwide. Injections in mice of whole Loxosceles (L.) intermedia venom or a recombinant toxin (rLiD1) produce systemic symptoms similar to those detected in envenomed humans.

Characterization of IgE-binding epitopes of peanut (Arachis hypogaea) PNA lectin allergen cross-reacting with other structurally related legume lectins.

Sera from peanut allergic patients contain IgE that specifically interact with the peanut lectin PNA and other closely related legume lectins like LcA from lentil, PsA from pea and PHA from kidney bean. The IgE-binding activity of PNA and legume lectins was assessed by immunoblotting, surface plasmon resonance (SPR) and ELISA measurements, using sera from peanut allergic patients as a IgE source. This IgE-binding cross-reactivity most probably depends on the occurrence of structurally related epitopes that have been identified on the molecular surface of PNA and other legume lectins.

In vivo protection against Tityus serrulatus scorpion venom by antibodies raised against a discontinuous synthetic epitope.

Scorpion stings cause human fatalities in numerous countries. Serotherapy is the only specific means to try to circumvent the noxious effects of venom toxins. TsNTxP is a natural anatoxin from the venom of the scorpion Tityus serrulatus that may be useful to raise therapeutic anti-venom sera.

Allergenicity of Hev b 13, a major esterase allergen in natural rubber latex (Hevea brasiliensis) allergy, does not only depend on its carbohydrate moiety.

The three-dimensional model built for the major latex allergen Hev b 13 consists of the typical organization of plant esterases made of a central bundle of five parallel beta-strands surrounded by five alpha-helices associated to two shorter alpha-helical segments. Up to 12 sets of sequential IgE-binding peptides were identified in SPOT experiments along the amino acid sequence of Hev b 13. They correspond in fact to eight IgE-binding epitopic stretches exposed on the surface of the allergen.

Identification of continuous interaction sites in PLA(2)-based protein complexes by peptide arrays.

Crotoxin (CA.CB) is a beta-neurotoxin from Crotalus durissus terrificus snake venom that is responsible for main envenomation effects upon biting by this snake. It is a heterodimer of an acidic protein (CA) devoid of any biological activity per se and a basic, enzymatically active, PLA(2) counterpart (CB).

Expression of Jug r 1, the 2S albumin allergen from walnut (Juglans regia), as a correctly folded and functional recombinant protein.

Jug r 1, the 2S albumin allergen from walnut, was isolated from ripe nuts as a native allergen and expressed in Escherichia coli using the Gateway technology as a recombinant allergen. The recombinant Jug r 1 (15 kDa) differs from the native allergen by the absence of cleavage of the polypeptide chain in two covalently associated light (3.5 kDa) and heavy (8 kDa) chains.

IgE-binding epitopic peptide mapping on a three-dimensional model built for the 13S globulin allergen of buckwheat (Fagopyrum esculentum).

The three-dimensional model built for the 13S globulin allergen of buckwheat (Fagopyrum esculentum) consists of three protomers exhibiting the cupin motif, arranged in a homotrimer around a three-fold symmetry axis. Using the SPOT technique, 11 continuous IgE-binding epitopic peptides were characterized on the molecular surface of the 13S globulin allergen of buckwheat. Except for one of them, they all correspond to well exposed regions containing electropositiveley and/or electronegatively charged residues, which cover up to 40% of the molecular surface of the allergen.

An in vivo protective response against toxic effects of the dermonecrotic protein from Loxosceles intermedia spider venom elicited by synthetic epitopes.

Loxoscelism is a necrotic-hemolytic syndrome caused by bites of brown spiders belonging to the genus Loxosceles. Many approaches for the treatment of Loxosceles poisoning have already been proposed, among which administration of specific antivenom is thought to be the more specific. We have evaluated the use of peptides as immunogen to raise in rabbits an antibody response that could protect animals from a challenge by the Loxtox isoform LiD1, one of the main toxic component of Loxosceles intermedia venom.

Mapping of IgE-binding epitopes on the major latex allergen Hev b 2 and the cross-reacting 1,3beta-glucanase fruit allergens as a molecular basis for the latex-fruit syndrome.

Nine distinct IgE-binding epitopes were identified along the entire amino acid sequence of the major latex allergen Hev b 2 (1,3beta-glucanase) using a set of synthetic 15-mer peptides frameshifted by 3 residues immobilized on cellulose membrane (Spot technique). Most of the amino acid residues building these IgE-binding epitopic regions are nicely exposed on the surface and the epitopes usually correspond to charged regions on the molecular surface of the protein. A smaller number of 5 IgE-binding epitopic areas was identified on the banana 1,3beta-glucanase, which exhibits a very similar overall conformation and charge distribution.

Characteristics, mechanisms of action, and epitope mapping of anti-factor VIII antibodies.

The development of anti-factor VIII (FVIII) antibodies (Abs), also called inhibitors, is currently one of the most serious complications arising during the treatment of hemophilia A patients. Improved prevention and eradication of these Abs remain a challenge both for clinicians and scientists. Numerous studies in the literature have reported on their epitope specificity, on their mechanism of FVIII inactivation, as well as on the methods used for their detection.

Mapping and conformational analysis of IgE-binding epitopic regions on the molecular surface of the major Ara h 3 legumin allergen of peanut (Arachis hypogaea).

Eight distinct sequential IgE-binding epitopes were identified along the amino acid sequence of Ara h 3 using the Spot technology. They essentially correspond to preferencially electropositive regions exposed on the molecular surface of the protein. A few IgE-binding epitopes are coalescent to create more extended IgE-binding regions exposed on the surface of the allergen.

A new method for 2D gel spot alignment: application to the analysis of large sample sets in clinical proteomics.

Background: In current comparative proteomics studies, the large number of images generated by 2D gels is currently compared using spot matching algorithms. Unfortunately, differences in gel migration and sample variability make efficient spot alignment very difficult to obtain, and, as consequence most of the software alignments return noisy gel matching which needs to be manually adjusted by the user.

Results: We present Sili2DGel an algorithm for automatic spot alignment that uses data from recursive gel matching and returns meaningful Spot Alignment Positions (SAP) for a given set of gels.

Simultaneous detection and epitope mapping of anti-factor VIII antibodies.

The development of antibodies (Abs) against infused factor VIII (FVIII) is currently one of the most serious complications in the treatment of patients suffering from haemophilia A. Improved prevention and eradication of these anti-FVIII Abs remain a challenge for both clinicians and scientists. Here we describe an immunoassay to simultaneously detect and map the epitope specificity of haemophilia A patients' inhibitors by screening plasma against both heavy and light chains (HC and LC) of human plasma-derived FVIII (pFVIII).

PEPOP: computational design of immunogenic peptides.

Background: Most methods available to predict protein epitopes are sequence based. There is a need for methods using 3D information for prediction of discontinuous epitopes and derived immunogenic peptides.

Results: PEPOP uses the 3D coordinates of a protein both to predict clusters of surface accessible segments that might correspond to epitopes and to design peptides to be used to raise antibodies that target the cognate antigen at specific sites.