Search PubMed⌕ Search

Biomedical subjects

L Otvos

Publications and source records attributed to L Otvos.

102 records · Page 6Linked to original sources
PubMed1989

Monoclonal antibodies to a synthetic peptide homologous with the first 28 amino acids of Alzheimer's disease beta-protein recognize amyloid and diverse glial and neuronal cell types in the central nervous system.

Studies were conducted to identify neural cells that synthesize and/or process cerebral amyloid using antisera and monoclonal antibodies (MAbs) raised to synthetic peptides based on the first 28 amino acids of the amyloid beta-protein. Using rabbit and mouse antisera, and 7 MAbs, sections of neocortex, hippocampus, cerebellum, and spinal cord from Alzheimer's disease (AD), Down's syndrome (DS), and control cases were probed. The antibodies produced 3 distinct immunohistochemical patterns: 1) staining restricted to neuritic plaque and blood vessel amyloid only (antisera, 1 of 7 MAbs); 2) immunoreactivity confined to cytoplasmic granules in diverse neuronal, glial (astrocytes, ependyma) and other (leptomeningeal, perivascular, choroid plexus) cells (1 of 7 MAbs); 3) a summation of these 2 patterns (5 of 7 MAbs). Controls resembled the AD and DS cases, except for a paucity of immunoreactive plaques and blood vessels in the controls. Immunoreactivity was reduced or removed by the peptides used to produce these antibodies. Formalin- and Bouins-fixed tissues reacted weakly or not at all with these antibodies while microwave denatured tissues reacted very intensely with them. Specific staining was enhanced by treatment of the tissue sections with Triton X-100, NaDodSO4, or trypsin. These studies significantly extend earlier studies that localized amyloid beta-protein precursor mRNA to human brain cells, and they suggest that the beta-protein, its precursor, and/or fragments thereof may exist in diverse neural cell types in AD, DS, and control brains.

Alzheimer Disease↗
PubMed1988

Recognition of hepatitis B surface antigen by human T lymphocytes. Proliferative and cytotoxic responses to a major antigenic determinant defined by synthetic peptides.

The antigenic sites for human T lymphocytes on hepatitis B surface Ag (HBsAg) were studied by using synthetic oligopeptides. T cell lines of the helper/inducer class, which were isolated from hepatitis B vaccine recipients, were found to react strongly and in an Ag-specific way with peptides corresponding to a sequence of 10 to 30 amino acids near the amino terminus of the HBsAg molecule. Cells with surface expression of the antigenic determinant contained in these synthetic peptides induced both proliferative and cytotoxic responses in the hepatitis B-specific T cells. The results indicate that amino acid residues 24-27 of HBsAg could be directly involved in this T cell determinant. Inhibition studies with mAb to MHC class II Ag and target cells from various HLA-typed individuals suggest that some T cell responses to this determinant of HBsAg might be restricted by the DPw4 molecule. However, the possibility exists that more than one of the MHC class II molecules could be involved as restricting elements of T cell responses to this synthetic peptide. In vivo experiments with synthetic peptides such as those described here are needed to demonstrate the possibility of enhancing HBsAg immune responses in some individuals.

Amino Acid Sequence↗
PubMed1988

Phosphorylation loops in synthetic peptides of the human neurofilament protein middle-sized subunit.

Peptides containing 13 and 39 amino acid residues and serine-side-chain-phosphorylated (P) analogues thereof, corresponding to human neurofilament protein middle-sized subunit (NF-M), have been synthesized in order to localize the phosphorylation site of this protein. The secondary structure of the nonphosphorylated peptides, determined by circular dichroism (CD) measurements, predicted secondary structural calculations and energy conformational calculations, was suggested to be a series of alternating type I (III). beta-turns and 3(10) or alpha-helices. By contrast, the phosphorylated peptides exhibit a unique conformation, probably due to salt bridges between the phosphoserine and the lysine residues. This has provided the first clear evidence that phosphorylation induces conformational changes among these synthetic peptides and presumably, in NF proteins as well. These phosphorylation loops might be the major recognition sites of the neurofilament protein-directed kinases.

Brain↗
PubMed1988

Alzheimer disease tangles share immunological similarities with multiphosphorylation repeats in the two large neurofilament proteins.

Immunological and structural analyses of neurofilament (NF) proteins with greater than 500 anti-NF monoclonal antibodies (mAbs) enumerated epitopes shared by NF proteins and Alzheimer neurofibrillary tangles. We identified the multiphosphorylation domain of the rat heaviest NF subunit--tandem repeats of Lys-Ser-Pro-Xaa (where Xaa is a small uncharged amino acid and serine is phosphorylated)--as the determinant recognized by 15 of the 16 mAbs from this collection of greater than 500 mAbs that detected neurofibrillary tangles. Most (11) of these 16 mAbs also recognized the previously characterized multiphosphorylation repeat in the human middle sized NF subunit. However, although these mAbs shared the ability to recognize NFTs, the antigen-binding domains of these 16 mAbs represented 13 separate classes based on their differential recognition of 12 synthetic peptides derived from the rat heaviest NF subunit and the human middle-sized NF subunit multiphosphorylation sites, NF subunits of 10 diverse species, and normal human tau protein. We conclude that NFTs share highly specific immunological and structural properties with specific rat heaviest NF subunit and human middle-sized NF subunit multiphosphorylation repeats.

Alzheimer Disease↗
PubMed1988

Identification of the major multiphosphorylation site in mammalian neurofilaments.

The sequence Lys-Ser-Pro-Val-Pro-Lys-Ser-Pro-Val-Glu-Glu-Lys-Gly repeats six times serially in the human midsized neurofilament (NF) protein (NF-M). To establish whether Lys-Ser-Pro-Val(Ala) is the major site for in vivo NF phosphorylation, peptides based on the human NF-M repeat were synthesized and chemically phosphorylated. These synthetic peptides were probed with 515 monoclonal antibodies (mAbs) that were raised to, and distinguished, several differentially phosphorylated forms of NF proteins. Studies with 95 of those mAbs that recognized the peptides before and after chemical phosphorylation demonstrated that a highly immunogenic epitope shared by the peptides is present in NFs from all species tested, including invertebrates. This suggests the phylogenetic conservation of a major NF phosphorylation site. Lastly, a cross-reactive antigenic determinant shared by the peptides and the major NF phosphorylation site was shown to exist in neurofibrillary tangles of patients with Alzheimer disease as well as in two neuron-specific microtubule-associated proteins (MAPs)--i.e., MAP2 and tau.

Animals↗
PubMed1988

Poly(amino2dA-dT) isomerizes into the unusual X-DNA double helix at physiological conditions inducing Z-DNA in poly (dG-methyl5dC).

It is demonstrated that a two-state conformational isomerization is induced in the poly(amino2-dA-dT) duplex by submillimolar concentrations of divalent magnesium cations in low-salt aqueous solution. The isomerization is fast and has a low degree of cooperativity. The resulting conformer is the unusual X-DNA double helix originally observed with poly(dA-dT) at very high concentrations of CsF. Interestingly, the X form is induced in poly(amino2dA-dT) under the physiological conditions when poly(dG-methyl5dC) assumes Z-DNA. The same conditions of stabilization are presumably connected with the fact, observed in previous phosphorus NMR studies, that Z- and X-DNA have similar polydinucleotide backbone architectures. Results presented in this work permit to specify base pair exocyclic groups responsible for the radically different conformational variability of the synthetic DNA molecules containing alternating purine-pyrimidine sequences of GC or AT base pairs.

Circular Dichroism↗
PubMed1987

Localization and immunological characterization of antigenic domains of the rabies virus internal N and NS proteins.

To locate epitopes on internal antigens of rabies virus, purified N and NS proteins of the nucleocapsid were cleaved at methionine, tryptophan or glutamic acid residues, transferred to nitrocellulose and immunostained using monoclonal antibodies (MAbs) specific for N and NS proteins, respectively. Five MAb-positive fragments of N protein and one fragment of NS protein were located after NH2-terminal amino acid sequence analysis within the deduced amino acid sequences of N and NS proteins. Antigenic analysis of synthetic overlapping peptides corresponding to the amino acid sequences of these fragments localized two major antigenic sites of N protein and one antigenic site of NS protein. Like the N- and NS-specific MAbs, anti-peptide antisera produced against the different synthetic antigens either reacted in a type-common fashion with all rabies virus strains, or in a type-specific manner with a restricted number of strains. The synthetic peptides corresponding to the three antigenic regions of the N and NS proteins also stimulated proliferation of human T lymphocytes derived from vaccinees who received inactivated rabies virus vaccine. This suggested that the antigenic regions of N and NS proteins are recognized by both B and T cells.

Amino Acid Sequence↗
PubMedDate not supplied

Is amyloid deposition in Alzheimer's disease preceded by an environment-induced double conformational transition?

Fragments of amyloid polypeptide (A4 or beta-protein) were synthesized on solid phase. Circular dichroism (CD) measurements showed that the N-terminal 1-12 fragment assumes an unordered conformation in water, but probably adopts a type I(III) beta-turn in trifluoroethanol (TFE). The central 11-25 fragment has a beta-sheet conformation in aqueous solution, while the full-length N-terminal 1-28 peptide shows helical features in TFE as well as in TFE-water mixtures. Based on secondary structural predictions, molecular mechanical calculations, and the differing and size-dependent conformational propensities of the smaller fragments in aqueous solution, the amyloid peptide is likely to undergo a double conformational transition that is characterized by a pleating process of its central segment. This conformational transition may start spontaneously above a critical peptide concentration, or it may be triggered by age-related or pathological changes ("watering") of the extracellular environment.

Alzheimer Disease↗
PubMedDate not supplied

Breakage in alpha-helix: a recognition site for anti-rabies virus ribonucleoprotein antibody.

Peptides composed of variants of a B-cell epitope followed by a T-cell determinant of rabies virus ribonucleoprotein (RNP) have been synthesized to elicit antibody production in mice. Conformation of the peptides was characterized by secondary structural prediction and circular dichroism measurements. It was found that only synthetic peptides with disrupted helical structure in the antigenic region were active on immunoblot assay, performed against a natural anti-protein monoclonal antibody (MAb) and provoked virus-neutralizing antibody production.

Amino Acid Sequence↗
PubMedDate not supplied

Coupling strategies in solid-phase synthesis of glycopeptides.

N-beta-(2-acetamido-2-deoxy-beta-D-glucopyranosyl)-asparaginyl peptides [Asn(GlcNAc)] corresponding to T-helper cell determinants were synthesized on solid-phase. Various amino-terminal- and carbohydrate-protecting groups were used on the glycosylated asparagine residue which was coupled to the peptide chain. We found that coupling rates decreased with increased size of the protected carbohydrate part of the acylating agent. Double couplings with an O-unprotected saccharide, as in Fmoc-Asn(GlcNAc)-OH resulted in acceptable coupling rates even with a synthetically difficult sequence corresponding to the T-cell epitopic peptide from the C-terminus of pigeon cytochrome c. The observed coupling rates on this peptide as well as on a T-cell epitopic pentapeptide, derived from the rabies virus N-protein, were comparable to those of conventional solid-phase peptide syntheses. The Fmoc-Asn(GlcNAc)-OH used can be prepared easily from commercially available components. The described glycopeptides will be used to probe effects of N-glycosylation on the immune recognition of viral glycoproteins.

Acetylglucosamine↗
PubMedDate not supplied

Solid phase synthesis and biological activity of rat relaxin.

The peptide hormone relaxin was isolated in good yield from the ovaries of the pregnant rodent Rattus rattus using a simplified purification schedule. It was subjected to comprehensive chemical characterization to confirm both its purity and predicted composition. The peptide was also chemically synthesized by the solid phase procedure. The two chains comprising the hormone were each assembled by the Boc-polystyrene method and, following conventional purification, combined in solution to form the single intramolecular and two intermolecular disulfide bonds. Following purification, the synthetic rat relaxin was fully chemically characterized and shown to be indistinguishable from the native peptide including by secondary structure analysis using circular dichroism spectroscopy. The native and synthetic rat relaxins were shown to be equally biologically active in the in vitro rat uterine relaxation assay and had pEC50 values that were comparable to synthetic human H2 relaxin.

Amino Acid Sequence↗