Atypical HIV serological profile of novel HIV-1 variant distinct from subtype O.
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AIDS Serodiagnosis↗
Biomedical subjects
R Deleys
Publications and source records attributed to R Deleys.
Design and production of a target-specific monoclonal antibody to parvovirus B19 capsid proteins.
Native parvovirus B19 was used as antigen to produce a mouse monoclonal antibody, R92F6, which reacted with B19 VP1 and VP2, neutralised the virus in bone marrow culture, and labelled infected cells in paraffin-embedded tissues from cases of B19-related fetal hydrops. The B19 epitope recognised by R92F6 (amino acids 328-344 from the amino terminal region of B19 VP2) appears to be highly conserved, since these tissue specimens were obtained over a 13 year period from widely spaced locations in the UK. This epitope was synthesised as a peptide (S7b) which was used as antigen to produce a mouse monoclonal antibody, 3H8, which specifically reacted with the B19 capsid proteins VP1 and VP2 in immunofluorescence and immunoblot assays. 3H8 was also capable of labelling formalin-fixed, paraffin-embedded, B19-infected fetal tissue and was shown to be of the same isotype as R92F6 (IgG1). Highly conserved epitopes derived from conserved amino acid sequences are valuable in the diagnosis of infectious disease. If these can be recognised and accurately synthesised, the production of specific mouse monoclonal antibodies may be possible for many human pathogens. Considering the vast amount of sequence data available in the literature, this approach seems to be both feasible and of wide potential.
Animals↗
Synthetic model peptides for apolipoproteins. I. Design and properties of synthetic model peptides for the amphipathic helices of the plasma apolipoproteins.
Amphipathic helical peptides are the lipid-binding motives of the plasma apolipoproteins, and synthetic peptide analogs have been used to unravel the mechanism of lipid association within this class of proteins. Hydrophobic interactions between the apolar amino acid residues belonging to the hydrophobic face of the amphipathic helices and the lipids are the major driving forces in the peptide-lipid association to form discoidal complexes. Ionic interactions and salt bridge formation between contiguous peptide chains in the complex can, however, contribute to the overall stability of the lipid-protein particle. This was studied by designing peptide analogs to the helical repeats of the apolipoproteins with variable degrees of salt bridge formation between adjacent peptide chains. The most stable conformation for pairs of synthetic peptides was calculated by energy minimisation together with the energy of interaction between peptides. The sequence of the peptides was derived from that of the 18A peptide synthesized by Segrest et al., and the theoretical calculations confirmed that ionic interactions between residues close to each other, along the edge of two adjacent anti-parallel peptides, can significantly contribute towards the stability of a peptide-phospholipid complex.
Amino Acid Sequence↗
Synthetic model peptides for apolipoproteins. II. Characterization of the discoidal complexes generated between phospholipids and synthetic model peptides for apolipoproteins.
The structure, composition and physico-chemical properties of complexes generated between phospholipids and synthetic model peptides for the amphipathic helices of the plasma apolipoproteins were studied. The sequences of the peptides were derived from that of the 18A peptide and designed to either enhance or decrease ionic interactions between pairs of peptides, as described in the accompanying paper. Complexes were prepared with dimyristoylphosphatidylcholine (DMPC), dipalmitoylphosphatidylcholine (DPPC), or with DPPC and cholesterol, and isolated on a Superose 6HR column. Association kinetics for the DMPC-peptides complexes were followed by measuring the turbidity as a function of the temperature. The diameters of the DPPC-peptide complexes, measured by gradient gel electrophoresis (GGE), were about 120 A. Fluorescence polarization measurements after labeling with diphenyl hexatriene (DPH) yielded transition temperatures of, respectively, 40.6, 41.5 and 41.8 degrees C for the DPPC/18AM1-, DPPC/18AM4- and DPPC/18A-peptide complexes. These values were confirmed by differential scanning calorimetry. Circular dichroism and infrared spectroscopy revealed that the peptides adopt an alpha-helical structure in solution and this percentage increased from 30-40% in the free peptides up to 50-60% in the complexes. Attenuated total reflection (ATR) infrared measurements of the complexes indicated that the peptides are oriented parallel to the acyl chains of the phospholipid bilayer. Denaturation of the peptides and of the peptide-lipid complexes was monitored by Trp fluorescence under addition of increasing amounts of GdmCl. The mid-points of the denaturation curves lie at, respectively, 0.05, 0.25 and 0.35 M GdmCl for the 18AM4, 18A and 18AM1 peptide and are shifted towards higher GdmCl concentrations after peptide-lipid binding. GdmCl denaturation decreased the alpha-helical content of the peptides and of the complexes, as monitored by circular dichroism measurement. The helix to random coil structure transition occurred at, respectively, 2.1, 2.2, and 2.0 M GdmCl for 18A, 18AM1 and 18AM4, compared to 5.1, 5.0, and 5.3 M in the corresponding complexes. These data suggest altogether that the structural properties, the mode of lipid-protein association and the stability of the phospholipid-peptide complexes are similar to those of native plasma apolipoproteins. The 18A and 18AM4 peptides which contain charged residues along the edge of the helix, leading to salt bridge formation between peptides were shown to mimic the amphipathic helices of the plasma apolipoproteins.
1,2-Dipalmitoylphosphatidylcholine↗
Phylogenetic analysis of gag genes from 70 international HIV-1 isolates provides evidence for multiple genotypes.
OBJECTIVE: To determine the extent of genetic variation among internationally collected HIV-1 isolates, to analyse phylogenetic relationships and the geographic distribution of different variants. DESIGN: Phylogenetic comparison of 70 HIV-1 isolates collected in 15 countries on four continents. METHODS: To sequence the complete gag genome of HIV-1 isolates, build multiple sequence alignments and construct phylogenetic trees using distance matrix methods and maximum parsimony algorithms. RESULTS: Phylogenetic tree analysis identified seven distinct genotypes. The seven genotypes were evident by both distance matrix methods and maximum parsimony analysis, and were strongly supported by bootstrap resampling of the data. The intra-genotypic gag distances averaged 7%, whereas the inter-genotypic distances averaged 14%. The geographic distribution of variants was complex. Some genotypes have apparently migrated to several continents and many areas harbor a mixture of genotypes. Related variants may cluster in certain areas, particularly isolates from a single city collected over a short time. CONCLUSIONS: The genetic variation among HIV-1 isolates is more extensive than previously appreciated. At least seven distinct HIV-1 genotypes can be identified. Diversification, migration and establishment of local, temporal 'blooms' of particular variants may all occur concomitantly.
Africa↗
HIV antigen detection in circulating immune complexes.
Various methods were evaluated for their effectiveness in releasing HIV antigen (Ag) from artificial immune complexes (IC) and from IC present in serum from HIV antibody (Ab) positive subjects. The most effective methods for recovering HIV Ag from IC were those which included a denaturation step to prevent reassociation of Ag with Ab. IC precipitation in 2.5% polyethylene glycol followed by acid treatment with 1 M glycine.HCl (pH 2) for 10 min at 70 degrees C in the presence of 0.05% SDS gave very satisfactory results. With this method, IC were detected in sera from HIV antibody positive Caucasian subjects at all stages of infection. After HIV IC dissociation, HIV Ag was detected in a significant number (8/17 or 47%) of asymptomatic subjects. IC were most prevalent during the late stages of infection. A substantial increase in HIV Ag positivity was also observed in 20 Senegalese HIV Ab positive sera. After HIV IC dissociation HIV antigen detection increased from 2/20 to 12/20. The relevance of IC detection is discussed.
Antigen-Antibody Complex↗