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Results for “HIV Core Protein p24”

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At least 19 recordsLinked to original sources
PubMed1996

Binding kinetics of an antibody against HIV p24 core protein measured with real-time biomolecular interaction analysis suggest a slow conformational change in antigen p24.

The interaction between HIV core protein p24 and the murine monoclonal antibody CB-4/1 or its Fab fragment showed unusual kinetics. Recombinant p24 was immobilised in a hydrophilic carboxymethyldextran matrix. At high concentration of CB-4/1 Fab the association of the antigen-antibody complex proceeds in two phases, while dissociation is mono-exponential. The antigen has a 'memory', i.e. shortly after dissociation of Fab-antigen complex the fast association phase is enhanced. Biphasic association was also found in solution. Experiments suggest a reversible change of binding properties in the epitope region with an overall time constant of about 100 s at room temperature. Intermediate steps with faster time constants must be involved. Slow conformational changes of p24 seem to be the most probable explanation. A simple model that provides a quantitative description of this process could not be found. Real-time analysis of antibody binding by surface plasmon resonance is a powerful method for studying such changes in the time domain of a few seconds to a few minutes.

Amino Acid Sequence↗
PubMed1991

Ozone inactivates HIV at noncytotoxic concentrations.

The inactivation of human immunodeficiency virus (HIV) and cytotoxic properties of ozone-treated serum and serum-supplemented media were examined. The titer of HIV suspensions in human serum was reduced in a dose-dependent manner when treated with total reacted ozone concentrations at a range of 0.5 to 3.5 micrograms/ml-1. Complete inactivation of HIV suspensions was achieved by 4.0 micrograms/ml-1 of ozone in the presence or absence of H-9 cells. In contrast, cellular metabolism, as measured by MTT dye cleavage, and DNA replication, as measured by BUdR incorporation, were enhanced in H-9 cells grown in media treated with quantities of ozone that completely inactivate HIV. The permissively HIV-infected cell line HXB/H-9 was cultured in ozone-treated media for six days with culture supernatants being sampled and assayed on alternate days for HIV p24 core protein. HIV p24 was reduced in all treated cultures compared to control cultures, with an average reduction of 46% [p24].

Blood↗
PubMed1991

Immune response to HIV p24 core protein during the early phases of human immunodeficiency virus infection.

The immune response to the p24 core antigen of human immunodeficiency virus type 1 (HIV-1) was studied in serial samples collected prospectively from 52 homosexual males in two separate cohorts from Amsterdam and San Francisco. p24 antibody levels were quantified with an antigen sandwich immunoassay using p24 recombinant antigen as capture and probe. Titers and slopes of dilution curves reflecting antibody affinity were analyzed. Only 45 of 52 men developed a measurable primary immune response to p24. In 17 (33%) patients there was a low response with maximum antibody titer below 66, shallow (low affinity) dilution curve, and 10 of the 17 became HIV antigen positive over a 2 year period. In 24 (46%) of the 52 patients titers ranged from 100-4000, steeper dilution curves were noted, and none became HIV antigen positive. Four (8%) men developed a strong immune response with high titers (greater than 12,000) and high affinity type dilution curve. Over time, after the peak immune response, antibody titer declined in some individuals related in part to the formation of immune complexes between HIV-1 p24 antigen and antibody which were dissociable. In vitro, the addition of increasing amounts of purified p24 antigen corresponded to decreasing antibody titer and a shallower dilution curve suggesting a preferential consumption of high affinity antibodies for complex formation. The magnitude of immune response to HIV-1 p24 antigen varies widely in infected homosexual men. Both the intrinsic ability to mount an immune response and immune complex formation contribute to the measurable antibody level.

Bisexuality↗
PubMed1991

The HIV core protein p24 inhibits interferon-gamma-induced increase of HLA-DR and cytochrome b heavy chain mRNA levels in the human monocyte-like cell line THP1.

Cells from the human monocytic cell-line THP1 were incubated prior to activation with IFN-gamma or LPS with varying amounts of p24, the main product of the HIV gag gene and the major component of the virus core. The IFN-gamma-dependent increase of mRNA for HLA-DR and for the heavy chain of cytochrome b was markedly decreased by p24 but not by gp120. This effect was abrogated by anti-p24 antibodies. On the other hand, preincubation of THP1 cells with p24 did not affect the accumulation of the LPS-dependent mRNA for TNF alpha and IL1-beta. These results indicate that p24 at concentrations similar to those found in the serum of HIV-infected individuals specifically affects IFN-gamma-induced activation markers.

Blotting, Northern↗
PubMed1994

Effects of tubeimoside-1 on HIV core protein p24 and cytopathogenesis in vitro.

Tubeimoside-1 (Tub) is a triterpenoid saponin first isolated in China from Bolbostemma paniculatum (Maxim) Franquet, Cucurbitaceae. To find out whether Tub has any anti-infective activity on human immunodeficiency virus (HIV), we tested its effects on HIV core protein p24 (with an ELISA) and HIV-mediated cytopathogenesis (using a colorimetric assay). The results showed that Tub inhibited both p24 production and cytopathogenesis mediated by HTLV-IIIB, and the median effective concentrations (EC50) were 24.1 and 22.9 micrograms.ml-1, respectively. Tub also effectively neutralized the infection of 2 other isolates, HTLV-IIIRF and HTLV-IIIMN. It is concluded that Tub had an inhibitory action on the infection of HIV-1 isolates and would be a promising candidate for treatment of AIDS.

Antiviral Agents↗
PubMed2000

Characterization of a discontinuous epitope of the human immunodeficiency virus (HIV) core protein p24 by epitope excision and differential chemical modification followed by mass spectrometric peptide mapping analysis.

A combination of epitope excision, epitope extraction, and differential chemical modification followed by mass spectrometric peptide mapping was used for the characterization of a discontinuous epitope that is recognized by the mouse anti-HIV-p24 monoclonal antibody 5E2.A3. In epitope excision, the protein is first bound to an immobilized antibody and then digested with proteolytic enzymes. In epitope extraction, the protein is first digested and subsequently allowed to react with the antibody. After epitope excision of the p24-5E2.A3 complex with endoproteinase Lys-C, a large fragment remained affinity bound corresponding to amino acids 1-158 of HIV-p24 (fragment 1-158). Further digestion, however, resulted in loss of affinity. Moreover, no affinity-bound fragments were observed after an epitope extraction experiment. These data from the epitope excision and extraction experiments suggest that the epitope is discontinuous. For the further characterization of the epitope, amino groups in the epitope-containing fragment were acetylated in both the affinity bound and free states followed by mass spectrometric analysis. Two successive acetylation reactions were performed: (1) the first used a low molar excess of acetic anhydride, and (2) the second, after separation from the antibody, a high molar excess of its hexadeuteroderivative. This isotopic labeling procedure, in combination with high resolution mass spectrometry, allowed the precise determination of relative reactivities of amino groups. In this study, no differences were observed in the ranking of the relative reactivities of five lysine residues. However, the N-terminal amino group was found to be part of the discontinuous epitope.

Acetylation↗
PubMed1995

Developmental and tissue-specific expression of human CD4 in transgenic rabbits.

A major obstacle to understanding AIDS is the lack of a suitable small animal model for studying HIV-1 infection and the subsequent development of AIDS, and for testing diagnostic, therapeutic, and preventive modalities. Our goal is to produce a rabbit model for the study of AIDS. Here we report on the generation of transgenic rabbits that express the human CD4 (hCD4) gene. The transgene, which contains the coding region for hCD4 and approximately 23 kb of sequence upstream of the translation start site, was used previously to direct hcD4 expression on the surface of CD4+ T cells of transgenic mice (Gillespie et al., 1993: Mol Cell Biol 13:2952-2958). The hCD4 transgene was detected in five males and two females derived from the microinjection in five males and two females derived from the microinjection of 271 rabbit embryos. Both hCD4 RNA and protein were expressed in peripheral blood lymphocytes (PBLs) from all five males but neither of the females. Human CD4 was expressed on PBLs from F1 offspring of all founder males. T-cell subset analysis revealed that hCD4 expression was restricted to rabbit CD4 (rCD4) expressing lymphocytes; mature rCD4- rCD8+ lymphocytes did not express hCD4. In preliminary studies, PBLs from hCD4 transgenic rabbits produced greater amounts of HIV-1 p24 core protein following HIV-1 infection in vitro than HIV-1 p24 antigen in nontransgenic rabbit infected cultures. These results extend to rabbits our previous observation that this transgene contains the sequence elements required for high-level expression in the appropriate cells of transgenic mice. Furthermore, these and previous studies demonstrating that expression of hCD4 protein enhances HIV-1 infection of rabbit T cells in vitro, coupled with reports that normal, nontransgenic rabbits are susceptible to HIV-1 infection, suggests that the hCD4 transgenic rabbits described herein will have an increased susceptibility to HIV-1 infection. In vivo HIV-1 infection studies with these rabbits are under way.

Animals↗
PubMed1998

Reactivity of sera from systemic lupus erythematosus and Sjögren's syndrome patients with peptides derived from human immunodeficiency virus p24 capsid antigen.

We have previously demonstrated that about one-third of patients with either Sjögren's syndrome (SS) or systemic lupus erythematosus (SLE) react to human immunodeficiency virus (HIV) p24 core protein antigen without any evidence of exposure to, or infection with, HIV itself. Herein, we further characterize the specificity of this reaction using enzyme-linked immunosorbent assay to peptides representing fragments of p24. Characteristic epitope-specific profiles were seen for SS and SLE patients. SS patients had significantly increased responses to peptides F (p24 amino acids 69 to 86) and H (amino acids 101 to 111) and diminished reactivity to peptides A (amino acids 1 to 16) and P (amino acids 214 to 228). SLE patients had increased reactivity to peptides E (amino acids 61 to 76), H, and P. Utilization of peptide P hyporeactivity as the criterion to select for SS patients results in a screen that is moderately sensitive (64%) and specific (79.3%). Adding hyperreactivity to one other peptide (F or H) as an additional criterion yields an expected decrease in sensitivity (to 41%) while increasing specificity (to 93.1%). All sera-reactive peptides from regions of known structure of HIV p24 were located in the apex of the p24 molecule. Thus, the specificity of the peptide reactivities described here indicates a specific pattern of a nonrandom cross-reactivity between HIV type 1 p24 and autoimmune sera which may be partially syndrome specific. The future focus of our work will be to optimize assays of the peptide as diagnostic tools.

Amino Acid Sequence↗
PubMed1992

Immunoglobulin V regions and epitope mapping of a murine monoclonal antibody against p24 core protein of HIV-1.

The nucleotide sequence of a murine monoclonal antibody (CB-mab-p24/13-5) against p24 core protein of the human immunodeficiency virus (HIV-1) was determined for variable regions of the heavy and light chain, respectively. Genetic elements encoding the VDJH- and VJL-regions of the antibody were generated from RNA by the polymerase chain reaction, cloned into the vector pICEM 19R and sequenced. Synthetic peptides, 10 amino acids overlapping served for the localization of the epitope. The residues 152-156 within the p24 sequence contain the epitope.

Amino Acid Sequence↗
PubMed1990

Peptide mapping on HIV polypeptides expressed in Escherichia coli. Quality control of different batches and identification of tryptic fragments containing residues of aromatic amino acids or cysteine.

Peptide mapping was used for the quality control of different batches of the recombinant HIV proteins p24 core and p24-gp41, expressed in Escherichia coli. These proteins comprise gag and env region polypeptides of the virus and may serve as suitable components in the diagnosis of HIV infections. The proteins were digested with trypsin and the mixtures were subjected to peptide mapping to prove batch equivalence of p24-gp41 and to isolate fragments of the p24-gp41 digest that differ from those of the p24 core digest. The proteins were reduced with dithiothreitol and the cysteine residues were derivatized by addition of 4-vinylpyridine. Peptide mapping was performed by means of reversed-phase high-performance liquid chromatography. Batch equivalence was proved by comparison of the maps. Peaks present in one map but not in the other were considered to be due to sequence differences or variability in digestion.

Amino Acid Sequence↗
PubMed1994

The ratio of defective HIV-1 particles to replication-competent infectious virions.

The ratio of infectious to defective particles has been investigated for human immunodeficiency virus type 1 (HIV-1). Although the concentration of HIV core p24 protein defined by ELISA permits the estimation of a total number of average-sized HIV particles, it does not provide information on the numerical value of infectious particles. This problem was addressed by limiting dilution of viral supernatant derived from various infected cell lines and then comparing the total number of HIV particles to the end-point titers of viral inoculum determined by reverse transcriptase assay and virus-induced cytotoxicity. The results indicate that the ratio of infectious to defective virus particles varies for different virus strains and host cells within a range from 1:1 to 1:100, but for given virus-cell system it remains constant.

Cell Line↗
PubMed1996

Lymphoid changes of the nasopharyngeal and palatine tonsils that are indicative of human immunodeficiency virus infection. A clinicopathologic study of 12 cases.

We report 12 cases in which the histomorphologic changes of the nasopharyngeal tonsils (adenoids) or palatine tonsils suggest infection with the human immunodeficiency virus (HIV). The patients included 10 men and two women, aged 20 to 42 years (median, 33 years). The clinical presentation included airway obstruction, pharyngitis, fever, and a tonsillar or adenoidal mass lesion. Histologic evaluation of the excised adenoids or tonsils in 10 of the cases demonstrated a spectrum of changes including florid follicular hyperplasia, follicle lysis, attenuated mantle zone, and the presence of multinucleated giant cells (MGC). The latter characteristically localized adjacent to the surface or tonsillar crypt epithelium. Two of the 12 cases showed marked lymphoid depletion with absent germinal centers, plasmacytosis, and stromal vascular proliferation. Immunohistochemical evaluation for HIV p24 core protein showed reactivity in 10 of 12 cases localized to follicular dendritic cell network (FDC), the MGC, scattered interfollicular lymphoid cells, and cells identified within the surface or crypt epithelium. Localization of viral RNA by in situ hybridization paralleled the HIV p24 immunohistochemical findings. Additional significant findings included the presence of both CD-68 and S-100 protein in the MGC and the presence of S-100 protein in dendritic cells. Other than HIV, no microorganisms were identified. At the time of presentation, eight patients were not known to be a risk for HIV infection, nor were they known to be HIV infected or suffering from AIDS. In these patients, HIV infection was suspected on the basis of the histologic changes seen in the resected tonsillar and adenoidal tissue. Serologic evaluation (by enzyme-linked immunosorbent assay), confirmed the presence of HIV infection. Our findings suggest the possibility of HIV dissemination through the upper aero-digestive tract mucosa via target cells, such as intraepithelial dendritic cells, submucosal macrophages, and T-lymphocytes. Subsequent presentation of viral antigens to the tonsillar and adenoidal lymphoid tissues results in enlargement of these structures that clinically may simulate a neoplastic proliferation but causes histomorphologic changes that are highly suspicious for HIV infection even in asymptomatic HIV-positive patients.

Adenoids↗
PubMed1991

Murine immune response to HIV-1 p24 core protein following subcutaneous, intraperitoneal and intravenous immunization.

The murine immune response to baculovirus-produced human immunodeficiency virus type-1 (HIV-1)p24 was examined after injection by three different routes: subcutaneously (s.c.), intraperitoneally (i.p.) and intravenously (i.v.). Both antigen-specific T-cell proliferation and serum antibody were induced by i.p. injection. In contrast, s.c. and i.v. injection of antigen resulted in specific antibody generation alone. Lympho-proliferative responses seen after i.p. injection were confined to splenocytes, and were greater after a low dose of antigen than after a high dose. p24-specific proliferation was not detected in lymph node cells. CD4:CD8 ratios were normal in lymph nodes and spleen at all times, irrespective of the dose or route of administration. p24-specific serum IgG antibodies were detected in all animals after the second injection of antigen. After s.c. and i.v. administration of high doses of antigen, the median antibody titres continued to rise after a third injection, but plateaued in animals injected by the i.p. route. In contrast, low doses of antigen given i.p. increased the median titre during and after the course of four injections. A low antigen dose given s.c. resulted in a plateau of median titre between the third and fourth injections. In i.v.-injected animals the median titre decreased between the third and fourth injections. IgG1 p24-specific antibodies were detected in all immunized mice, whereas IgM antibodies were detectable only following i.p. injection of antigen.

Animals↗
PubMed1993

Monoclonal antibodies to HIV-1 p24 core protein include pairs which exhibit synergistic binding.

Gamma and kappa chain cDNAs from four mouse monoclonal antibodies (mAbs) which bind three different sites on the core antigen (p24) of HIV-1 have been cloned and their V-region sequences determined. These mAbs are part of a larger group of seven anti-p24 mAbs analyzed in simultaneous competition assays with HIV-1 lysate as antigen and in protein blotting experiments using 10 carboxy-terminal truncations of a p24 fusion protein. One mAb, BB128, recognizes the p24 loop sequence EAAEWDRVHP and enhances the binding of two other mAbs (BI1777 and BI1279) when tested pairwise in simultaneous competition assays. The two monoclonals enhanced by BB128 recognize different antigenic sites on p24, with BI1777 binding to carboxy-terminal sequences and BI1279 to amino-terminal residues. In the pairwise assays, mAb BI1279 also acts as enhancing antibody for BI1777, as does mAb BB328, which recognizes residues in the central region of p24. Since aggregated p24 monomers form the HIV-1 capsid, p24 is a multivalent antigen in HIV-1 lysate. It seems likely, therefore, that synergistic binding of mAb pairs to p24 is effected by bivalent binding of the enhancing mAb stabilizing a conformation favorable for bivalent binding of the enhanced mAb.

Amino Acid Sequence↗
PubMed1991

Detection of p24 in HIV-1 infected cells embedded in LR White and Lowicryl K4M.

In this study we present a postembedding on-grid immunogold labelling procedure for the ultrastructural localization of the HIV-1 core protein p24. HIV-1 infected cells were fixed in 0.1% glutaraldehyde, incompletely dehydrated and embedded in LR White or in Lowicryl K4M. Antigenic sites were detected by incubation of ultrathin sections with primary mouse monoclonal antibody anti-HIV-1 p24, followed by the secondary antibody goat anti-mouse IgG coupled to 10nm gold particles. Antigenicity of p24 was found to withstand the applied fixation and was shown to be preserved in LR White as well as in Lowicryl. The described procedure permits the uncomplicated and easy detection of p24 in HIV-1 infected cells and tissues.

Acrylic Resins↗
PubMed1993

B-cell abnormalities in AIDS: stable and clonally-restricted antibody response in HIV-1 infection.

Antibodies in sera of HIV-1 infected individuals against the HIV-1 core protein (p24), HIV-1 envelope glycoprotein (gp120) and reverse transcriptase (RT) are characterized by a skewed light chain isotype expression. The kappa/lambda ratios of antibodies to p24 and gp120 in infected individuals were found to be unique in each individual, but constant over several years independently from disease progression. The oligoclonal nature of the anti-HIV-1 antibodies suggested by skewed kappa/lambda expression was confirmed with isoelectric focusing of affinity purified antibodies to p24 and gp120. Analysis of the utilization of V gene families in purified anti-p24 and anti-gp120 antibodies revealed a restricted and biased VH gene family usage. In contrast, the utilization of VK gene families appeared to be random. The finding of stable and restricted antibody responses in infected individuals could be one of the causes for the failure to produce antibodies to HIV-1 that are effective against escape virus variants.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Human immunodeficiency virus type 1 p24 production and antigenic variation in tissue culture of isolates with various growth characteristics.

Two monoclonal antibodies (Mabs) reacting with different epitopes of the human immunodeficiency virus type 1 core protein p24 (HIV p24) were used either singly or in combination as tracers in enzyme-linked immunosorbent assays. The culture supernatant of 215 samples of peripheral blood mononuclear cells from 112 patients were measured for HIV p24 and reverse transcriptase (RT) activity during cultivation. One hundred forty-one cultures were positive for HIV p24 and 122 for RT after 32 days of cultivation. After 8-9 days, HIV p24 was detected in 50.4% and RT in 27.8% of the cultures later judged as HIV positive. Two patients seemed to have substrains of HIV-1 not reactive with one of the Mabs.

Acquired Immunodeficiency Syndrome↗
PubMed1994

Defective infection of rabbit peripheral blood monocyte cultures with human immunodeficiency virus type 1.

Human immunodeficiency virus type 1 (HIV-1) produces abortive infections of primary cultures of rabbit peripheral blood mononuclear cells (PMBCs). Mitogen activation of rabbit PBMCs or the addition of exogenous cytokines to the cultures does not change the level of release of the early HIV core protein, p24, into the culture medium. The amount of p24 increases steadily and reaches peak levels about 7 days after infection. HIV-1-specific DNA env sequences are also detected in infected PBMCs. However, reverse transcription of RNA of samples from activated infected cultures to cDNA, followed by amplification by the polymerase chain reaction, revealed transcription of gag, but not env, regions, suggesting that HIV-1 infection of rabbit PBMC does not lead to the replication and maturation of complete HIV-1 virions. In addition, neither CPE nor lytic infection was observed in HIV-1-infected rabbit cells and infectivity could not be transferred from rabbit cells infected with HIV for up to 2 weeks to MT-2 or H9 indicator cell lines. In addition, no CPE was seen in long-term cultures of the HTLV-I-transformed rabbit cell line PLT-1441, after inoculation with HIV. It is concluded that primary rabbit PBMCs may be infectable by HIV-1 but are not permissive for production of infectious virus. This conclusion is consistent with the apparent long-term latent infection seen after inoculation of rabbits with HIV-1 or HIV-1-infected cells.

Animals↗