André Tartar

Multiepitopic B- and T-Cell Responses Induced in Humans by a Human Immunodeficiency Virus Type 1 Lipopeptide Vaccine

ABSTRACT We have attempted to develop an anti-human immunodeficiency virus (HIV) lipopeptide vaccine with several HIV-specific long peptides modified by C-terminal addition of a single palmitoyl chain. A mixture of six lipopeptides derived from regulatory or structural HIV-1 proteins (Nef, Gag, and Env) was prepared. A phase I study was conducted to evaluate immunogenicity and tolerance in lipopeptide vaccination of HIV-1-seronegative volunteers given three injections of either 100, 250, or 500 μg of each lipopeptide, with or without immunoadjuvant (QS21). This report analyzes in detail B- and T-cell responses induced by vaccination. The lipopeptide vaccine elicited strong and multiepitopic B- and T-cell responses. Vaccinated subjects produced specific immunoglobulin G antibodies that recognized the Nef and Gag proteins. After the third injection, helper CD4+-T-cell responses as well as specific cytotoxic CD8+ T cells were also obtained. These CD8+ T cells were able to recognize naturally processed viral proteins. Finally, specific gamma interferon-secreting CD8+ T cells were also detected ex vivo.

Parallel synthesis of 1,2,4-oxadiazoles from carboxylic acids using an improved, uronium-based, activation

Abstract We describe the synthesis of 1,2,4-oxadiazoles from carboxylic acids and amidoximes using 2-(1 H -benzotriazole-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU) as an activating agent of the carboxylic acid function for the O -acylation step. This method was used for the synthesis of a library of 24 1,2,4-oxadiazoles.

Lipopeptides induce cell-mediated anti-HIV immune responses in seronegative volunteers.

ObjectiveTest the efficacy of a mixture of six NEF (N1, N2, N3), GAG (G1, G2) and ENV (E) lipopeptides in the induction of B- and T-cell anti-HIV responses. DesignA randomized phase I open-label dose-finding trial. Twenty-eight healthy seronegative volunteers received the lipopeptides, with or without the adjuvant QS21. MethodsAnti-HIV-peptide antibodies were detected by enzyme-linked immunosorbent assay and Western blotting. Induction of cellulary responses was assessed by proliferative test and 51Cr-release assay. ResultsLocal and systemic adverse reactions were always mild or moderate. After three injections an antibody response was detected in 25 out of 28 volunteers (89%). T cells from 19 (79%) of the 24 volunteers proliferated in response to at least one peptide. The majority of the volunteers had induced a multispecific proliferative response; that is, cells from volunteers proliferated to two (five of 19), three (five of 19), four (three of 19) or five peptides (one of 19). Cytotoxic responses by anti-HIV CD8+ lymphocytes could be tested in 24 volunteers, 13 (54%) of whom had clear and reproducible responses, with strong activity in the remaining 12 (> 20% of specific lysis), and polyepitopic responses were detected in at least seven of the 13 responders. Cytotoxic responses were found against the whole NEF protein (clade B LAI) in three of four tested volunteers and cross-reactions with the proteins of clade B (MN) and clade A (Bangui) HIV-1 strains, and also HIV-2 ROD, were detected in one of two tested volunteers. ConclusionsLipopeptides are promising immunogens for an AIDS vaccine.

The complete amino acid sequence of bovine milk angiogenin

The amino acid sequence of angiogenin isolated from bovine milk was deduced by gas‐phase sequencing of the protein and its fragments. The protein contains 125 residues and has a calculated molecular mass of 14 577 Da. The sequence is highly homologous (65% identity) to the sequence of human angiogenin, most of the differences being the result of conservative replacements. Like human angiogenin, the bovine protein is also homologous to bovine pancreatic RNase A (34% identity) and the three major active site residues known to be involved in the catalytic process, His‐14, Lys‐41 and His‐115, are conserved. When tested against conventional substrates for RNase A activity, bovine angiogenin displays the same selective ribonucleolytic activity as human angiogenin. The sequence of bovine angiogenin contains the cell recognition tripeptide Arg‐Gly‐Asp which is not present in the human protein.

Slow isomerization of some proline-containing peptides inducing peak splitting during reversed-phase high-performance liquid chromatography

Abstract A slow conformational equilibrium, commensurable with the retention times, was shown to induce peak broadening or peak splitting during reversed-phase high-performance liquid chromatography of several medium-sized peptides. Elution at 50°C resulted in sharp unique peaks, while at sub-ambient temperature well resolved peaks were observed. Linear peptides which show this phenomenon had a Pro-Pro bond, but the phenomenon was also observed in the case of a cyclic peptide containing two non-vicinal proline residues.

Intramolecular mapping of Plasmodium falciparum P126 proteolytic fragments by N-terminal amino acid sequencing.

Protein P126 (also called P140, P113, SERA, SERP1) is a major parasitophorous vacuole antigen of Plasmodium falciparum. This protein is processed upon merozoite release into 2 fragments of 73 kDa (P73) and 50 kDa (P50), which are found in the culture medium. P73 is composed of 2 polypeptides of 47 and 18 kDa linked by disulfide bridges. In the presence of leupeptin, an inhibitor of serine and cysteine proteases which inhibits merozoite release, a 56-kDa intermediate product (P56) is recovered in the culture medium instead of P50. In order to map these proteolytic fragments on the 126-kDa precursor, we purified them from Plasmodium falciparum culture medium by immunoadsorption, SDS-electrophoresis and Western blotting on PVDF membrane and determined the N termini of P126, P73 (P47 and P18), P50 and P56. Comparison of these sequences with the amino acid sequence deduced from the P126 gene allowed the mapping of the different fragments on the precursor. P47 was at the N-terminal and P18 at the C-terminal end of P126. P56 and P50 had the same N-termini and were located in the middle of P126. This latter result indicates that the proteolysis of P56-P50 occurs at the C-terminus of P56. The peptide bonds cleaved by leupeptin-insensitive activities are Glu-Thr and Gln-Asp; C-terminal sequencing of P50 will be needed to identify the leupeptin-sensitive cleavage site.

A novel merozoite surface antigen of Plasmodium falciparum (MSP-3) identified by cellular-antibody cooperative mechanism antigenicity and biological activity of antibodies

We report the identification of a 48kDa antigen targeted by antibodies which inhibit Plasmodium falciparum in vitro growth by cooperation with blood monocytes in an ADCI assay correlated to the naturally acquired protection. This protein is located on the surface of the merozoite stage of P. falciparum, and is detectable in all isolates tested. Epidemiological studies demonstrated that peptides derived from the amino acid sequence of MSP-3 contain potent B and T-cell epitopes recognized by a majority of individuals living in endemic areas. Moreover human antibodies either purified on the recombinant protein, or on the synthetic peptide MSP-3b, as well as antibodies raised in mice, were all found to promote parasite killing mediated by monocytes.

Parallel synthesis of polysubstituted tetrahydroquinolines

Abstract A three-component cycloaddition was used to prepare a library of polysubtituted tetrahydroquinolines. Reaction conditions were optimised and a large range of anilines, aldehydes and alkenes were tested.

Characterization and synthesis of a macrophage inhibitory peptide from the second constant domain of human immunoglobulin G

We have shown that IgG hydrolysed by Schistosoma mansoni schistosomula inhibited various macrophage functions, especially phagocytosis and anti‐schistosome cytotoxicity. Here we show that a tripeptide, Thr289‐Lys‐Pro291, of the second constant domain of human immunoglobulin G (peptide 286–292) reproduced the inhibitory effect of a total hydrolysate. Indeed the β‐glucuronidase release from IgE‐anti‐IgE‐stimulated rat and human macrophages decreased and its intracellular level did not rise after a prior incubation of the cells with Thr‐Lys‐Pro (500 nmol/ml). Moreover, the cell migration as well as the superoxide anion O− 2 generation were 50–80% reduced by the tripeptide. These results suggest that a single peptide set may be responsible for the decrease of the macrophage functions at the early stage of the parasite infection in the mammalian host. The pharmacologic properties of this tripeptide are under investigation.

Solution conformation of leiurotoxin I (scyllatoxin) by 1H nuclear magnetic resonance. Resonance assignment and secondary structure.

A proton NMR study at 500 MHz of leiurotoxin I in water is presented. Nearly complete sequence‐specific assignments of the individual backbone and side‐chain proton resonances were achieved using through‐bond and through‐space connectivities obtained from standard two‐dimensional NMR techniques. The secondary structure of this toxin is inferred from a combination of short‐range nuclear Overhauser enhancements, scalar couplings and proton/deuteron exchange rates. Three disulflde bridges locate the N‐terminal part (that is α‐helical from residue 6 to 16) on one side of a C‐terminal two stranded antiparallel β sheet (from Leu18 to Val29). The latter features a tight turn at Gly23‐Asp24.