scholarly journals Human Immunodeficiency Virus-Infected Patients with Prior Pneumocystis Pneumonia Exhibit Increased Serologic Reactivity to Several Major Surface Glycoprotein Clones

2006 ◽  
Vol 13 (10) ◽  
pp. 1071-1078 ◽  
Author(s):  
K. R. Daly ◽  
J. V. Koch ◽  
N. J. Shire ◽  
L. Levin ◽  
P. D. Walzer
Keyword(s):  
Human Immunodeficiency Virus ◽  
Blood Donors ◽  
The Other ◽  
Surface Glycoprotein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Immune Reactivity ◽  
Major Surface Glycoprotein ◽  
Immunodeficiency Virus ◽  
Major Surface ◽  
Hiv 1

ABSTRACT Recombinant clones of the carboxyl terminus of the major surface glycoprotein (MsgC) of Pneumocystis jirovecii are useful for analyzing serologic responses in humans. However, there is no standardized set of antigens in general use, which could lead to conflicting results. We have previously shown that human immunodeficiency virus type 1 (HIV-1)-infected patients with prior Pneumocystis pneumonia (PcP+) responded more frequently and more strongly to a clone of MsgC than did HIV-1-infected patients without PcP (PcP−). Here we test three new clones of MsgC to determine the effect of antigenic sequence variation on immune reactivity in blood donors and HIV-infected patients previously analyzed for reactivity to our original MsgC clone. In Western blot analyses, PcP+ patients exhibited the highest frequency of reactivity to each MsgC clone, and the frequency of reactivity with all four MsgC clones together was significantly higher in sera from PcP+ patients than in sera from the other patient groups. Furthermore, in an enzyme-linked immunosorbent assay we found that the PcP+ population had the highest level of reactivity to two of the four clones tested. One of the new clones could distinguish between PcP+ and PcP− populations, and two MsgC clones could distinguish blood donors from the other patient populations. The results show that inherent differences in MsgC amino acid sequence can affect recognition by antibodies independently of variations in protein length or patient population, and the utility of a clone depends on its sequence and on the populations tested.

scholarly journals Analisis filogenetik gen mitochondrial large subunit (mtLSU) Pneumocystis jirovecii pada Pasien Human Immunodeficiency Virus (HIV) Terduga Pneumoniae di Jakarta

2020 ◽  
Vol 9 (1) ◽  
pp. 9-18
Author(s):  
Conny Riana Tjampakasari ◽  
Andi Yasmon ◽  
Agus Sjahrurachman ◽  
Samsuridjal Djauzi
Keyword(s):  
Human Immunodeficiency Virus ◽  
Hot Spot ◽  
Large Subunit ◽  
Surface Glycoprotein ◽  
Pneumocystis Jirovecii ◽  
Rt Pcr ◽  
Major Surface Glycoprotein ◽  
Immunodeficiency Virus ◽  
Polymerase Chain ◽  
Major Surface

Abstrak Pneumocystis jirovecii (P. jirovecii) merupakan patogen oportunistik yang penting pada pasien dengan gangguan kekebalan menurun khususnya human immunodeficiency virus (HIV). P. jirovecii tersebar dimanamana, menyebar melalui udara, dan menyerang sistem pernapasan atas. P. jirovecii mempunyai beberapa faktor virulensi, antara lain major surface glycoprotein (MSG) yang merupakan antigen yang paling banyak ditemukan di permukaan. Pendekatan biologi molekuler digunakan untuk mempelajari patogen ini karena hingga saat ini kultur belum dapat dilakukan. Penelitian ini bertujuan untuk memperoleh data genotip yang dapat dimanfaatkan sebagai dasar data demografi dan epidemiologi molekuler P. jirovecii di Indonesia. Dua puluh sampel sputum positif P. jirovecii pada real-time polymerase chain reaction (RT-PCR) dilakukan karakterisasi terhadap gen mitochondrial large subunit (mtLSU). Virulensi daerah hot spot gen mtLSU dianalisis dengan metode PCR dan sekuensing deoxyribonucleic acid (DNA). Diperoleh 30 strain dengan 7 varian didominasi oleh varian 3 yang bersirkulasi di Jakarta. Analisis filogenetik dengan strain negara lain menunjukkan strain Jakarta berkerabat dekat dengan strain Iran, India dan Korea. Kata kunci : Pneumocystis jirovecii, mtLSU, PCR, filogenetik Abstract Pneumocystis jirovecii (P. jirovecii) is an important opportunistic pathogen in immunocompromised patients especially human immunodeficiency virus (HIV). P. jirovecii is spread everywhere, spread through the air, and attacking the upper respiratory system. P. jirovecii has several virulence factors, including major surface glycoprotein (MSG) which is the most widely found on the surface antigen. The molecular biology approach is used to study this pathogen because until now culture cannot be done. This study aims to obtain genotype data that can be used as a basis for demographic and molecular epidemiological data of P. jirovecii in Indonesia. Twenty P. jirovecii positive sputum samples on real-time polymerase chain reaction (RT-PCR) were characterized by the mitochondrial large subunit (mtLSU) gene. Virulence of the mtLSU gene hot spot region was analyzed by PCR method and deoxyribonucleic acid (DNA) sequencing. Obtained 30 strains with 7 variants dominated by variant 3 circulating in Jakarta. Phylogenetic analyzed with strains of other countries shows that Jakarta strains are closely related to strains of Iran, India dan Korea. Keywords: Pneumocystis jirovecii, mtLSU, PCR, phylogenetic

scholarly journals Amino-Terminal Substitutions in the CCR5 Coreceptor Impair gp120 Binding and Human Immunodeficiency Virus Type 1 Entry

1998 ◽  
Vol 72 (1) ◽  
pp. 279-285 ◽  
Author(s):  
Tatjana Dragic ◽  
Alexandra Trkola ◽  
Steven W. Lin ◽  
Kirsten A. Nagashima ◽  
Francis Kajumo ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Surface Glycoprotein ◽  
Alanine Scanning Mutagenesis ◽  
Cc Chemokine ◽  
Amino Terminal ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The CC-chemokine receptor CCR5 is required for the efficient fusion of macrophage (M)-tropic human immunodeficiency virus type 1 (HIV-1) strains with the plasma membrane of CD4+ cells and interacts directly with the viral surface glycoprotein gp120. Although receptor chimera studies have provided useful information, the domains of CCR5 that function for HIV-1 entry, including the site of gp120 interaction, have not been unambiguously identified. Here, we use site-directed, alanine-scanning mutagenesis of CCR5 to show that substitutions of the negatively charged aspartic acid residues at positions 2 and 11 (D2A and D11A) and a glutamic acid residue at position 18 (E18A), individually or in combination, impair or abolish CCR5-mediated HIV-1 entry for the ADA and JR-FL M-tropic strains and the DH123 dual-tropic strain. These mutations also impair Env-mediated membrane fusion and the gp120-CCR5 interaction. Of these three residues, only D11 is necessary for CC-chemokine-mediated inhibition of HIV-1 entry, which is, however, also dependent on other extracellular CCR5 residues. Thus, the gp120 and CC-chemokine binding sites on CCR5 are only partially overlapping, and the former site requires negatively charged residues in the amino-terminal CCR5 domain.

scholarly journals Structure-Function Analysis of the Epitope for 4E10, a Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody

2006 ◽  
Vol 80 (4) ◽  
pp. 1680-1687 ◽  
Author(s):  
Florence M. Brunel ◽  
Michael B. Zwick ◽  
Rosa M. F. Cardoso ◽  
Josh D. Nelson ◽  
Ian A. Wilson ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Function Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Peptide Epitope ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
4E10 Antibody

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) neutralizing antibody 4E10 binds to a linear, highly conserved epitope within the membrane-proximal external region of the HIV-1 envelope glycoprotein gp41. We have delineated the peptide epitope of the broadly neutralizing 4E10 antibody to gp41 residues 671 to 683, using peptides with different lengths encompassing the previously suggested core epitope (NWFDIT). Peptide binding to the 4E10 antibody was assessed by competition enzyme-linked immunosorbent assay, and the Kd values of selected peptides were determined using surface plasmon resonance. An Ala scan of the epitope indicated that several residues, W672, F673, and T676, are essential (>1,000-fold decrease in binding upon replacement with alanine) for 4E10 recognition. In addition, five other residues, N671, D674, I675, W680, and L679, make significant contributions to 4E10 binding. In general, the Ala scan results agree well with the recently reported crystal structure of 4E10 in complex with a 13-mer peptide and with our circular dichroism analyses. Neutralization competition assays confirmed that the peptide NWFDITNWLWYIKKKK-NH2 could effectively inhibit 4E10 neutralization. Finally, to limit the conformational flexibility of the peptides, helix-promoting 2-aminoisobutyric acid residues and helix-inducing tethers were incorporated. Several peptides have significantly improved affinity (>1,000-fold) over the starting peptide and, when used as immunogens, may be more likely to elicit 4E10-like neutralizing antibodies. Hence, this study represents the first stage toward iterative development of a vaccine based on the 4E10 epitope.

scholarly journals Immunoglobulin G3 from Polyclonal Human Immunodeficiency Virus (HIV) Immune Globulin Is More Potent than Other Subclasses in Neutralizing HIV Type 1

2001 ◽  
Vol 75 (14) ◽  
pp. 6558-6565 ◽  
Author(s):  
Orit Scharf ◽  
Hana Golding ◽  
Lisa R. King ◽  
Nancy Eller ◽  
Doug Frazier ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Immune Globulin ◽  
High Titer ◽  
Passive Immunization ◽  
Hinge Region ◽  
Enzyme Linked Immunosorbent Assay ◽  
Fab Fragments ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Passive antibody prophylaxis against human immunodeficiency virus type 1 (HIV-1) has been accomplished in primates, suggesting that this strategy may prove useful in humans. While antibody specificity is crucial for neutralization, other antibody characteristics, such as subclass, have not been explored. Our objective was to compare the efficiencies of immunoglobulin G (IgG) subclasses from polyclonal human HIV immune globulin (HIVIG) in the neutralization of HIV-1 strains differing in coreceptor tropism. IgG1, IgG2, and IgG3 were enriched from HIVIG by using protein A-Sepharose. All three subclasses bound major HIV-1 proteins, as shown by Western blot assay and enzyme-linked immunosorbent assay. In HIV-1 fusion assays using X4, R5, or X4R5 envelope-expressing effector cells, IgG3 more efficiently blocked fusion. In neutralization assays with cell-free viruses using X4 (LAI, IIIB), R5 (BaL), and X4R5 (DH123), a similar hierarchy of neutralization was found: IgG3 > IgG1 > IgG2. IgG3 has a longer, more flexible hinge region than the other subclasses. To test whether this is important, IgG1 and IgG3 were digested with pepsin to generate F(ab′)2 fragments or with papain to generate Fab fragments. IgG3 F(ab′)2 fragments were still more efficient in neutralization than F(ab′)2 of IgG1. However, Fab fragments of IgG3 and IgG1 demonstrated equivalent neutralization capacities and the IgG3 advantage was lost. These results suggest that the IgG3 hinge region confers enhanced HIV-neutralizing ability. Enrichment and stabilization of IgG3 may therefore lead to improved HIVIG preparations. The results of this study have implications for the improvement of passive immunization with polyclonal or monoclonal antibodies and suggest that HIV-1 vaccines which induce high-titer IgG3 responses could be advantageous.

scholarly journals Characterization of a Major Histocompatibility Complex Class II X-Box-Binding Protein Enhancing Tat-Induced Transcription Directed by the Human Immunodeficiency Virus Type 1 Long Terminal Repeat

2000 ◽  
Vol 74 (19) ◽  
pp. 8989-9001 ◽  
Author(s):  
Carlo Mischiati ◽  
Giordana Feriotto ◽  
Monica Borgatti ◽  
Patrizio Giacomini ◽  
Roberto Gambari
Keyword(s):  
Human Immunodeficiency Virus ◽  
Transcription Factors ◽  
Major Histocompatibility Complex ◽  
Class Ii ◽  
The Other ◽  
Nuclear Proteins ◽  
Histocompatibility Complex ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The X-box element present within the promoter region of genes belonging to the major histocompatibility complex (MHC) plays a pivotal role in the expression of class II molecules, since it contains the binding sites for several well-characterized transcription factors. We have analyzed a randomly selected compilation of viral genomes for the presence of elements homologous to the X box of the HLA-DRA gene. We found that human immunodeficiency virus type 1 (HIV-1) shows the highest frequency of X-like box elements per 1,000 bases of genome. Within the HIV-1 genome, we found an X-like motif in the TAR region of the HIV-1 long terminal repeat (LTR), a regulative region playing a pivotal role in Tat-induced HIV-1 transcription. The use of a decoy approach for nuclear proteins binding to this element, namely, XMAS (X-like motif activator sequence), performed by transfection of multiple copies of this sequence into cells carrying an integrated LTR-chloramphenicol acetyltransferase construct, suggests that this element binds to nuclear proteins that enhance Tat-induced transcription. In this report we have characterized two proteins, one binding to the XMAS motif and the other to the flanking regions of XMAS. Mobility shift assays performed on crude nuclear extracts or enriched fractions suggest that similar proteins bind to XMAS from HIV-1 and the X box of the HLA-DRA gene. Furthermore, a UV cross-linking assay suggests that one protein of 47 kDa, termed FAX (factor associated with XMAS)-1, binds to the XMAS of HIV-1. The other protein of 56 kDa was termed FAX-2. In a decoy ex vivo experiment, it was found that sequences recognizing both proteins are required to inhibit Tat-induced HIV-1 LTR-driven transcription. Taken together, the data reported in this paper suggest that XMAS and nearby sequences modulate Tat-induced HIV-1 transcription by binding to the X-box-binding proteins FAX-1 and FAX-2. The sequence homology between XMAS and X box is reflected in binding of a common protein, FAX-1, and similar functional roles in gene expression. To our knowledge, this is the first report showing that transcription factors binding to the X box of the MHC class II genes enhance the transcription of HIV-1.

scholarly journals Antiphosphatidylserine antibodies in human immunodeficiency virus-1 patients with evidence of T-cell apoptosis and mediate antibody- dependent cellular cytotoxicity [see comments]

Blood ◽  
1996 ◽  
Vol 87 (12) ◽  
pp. 5185-5195 ◽  
Author(s):  
F Silvestris ◽  
MA Frassanito ◽  
P Cafforio ◽  
D Potenza ◽  
M Di Loreto ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cell ◽  
Cell Apoptosis ◽  
Clinical Feature ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
T Cell Apoptosis ◽  
Immunodeficiency Virus ◽  
Hiv 1

Serum reactivities to a panel of phospholipid antigens, including cardiolipin (CL), phosphatidylserine (PS), sphingomyelin, phosphatidylcholine, and phosphatidylethanolamine, were measured by enzyme-linked immunosorbent assay in 196 human immunodeficiency virus- l+ (HIV-1+) patients with CDC II to IVC clinical disease. Significant levels of IgG to CL, PS, or both were observed in 23 patients lacking evidence of thrombophilic events or any peculiar clinical feature of HIV-1 infection. Fluorescence-activated cell sorting analyses showed that in vitro apoptosis of T cells was increased in patients with high serum anti-PS IgG, whereas the overexpression of Fas/Apo-1 marker was detected in all patients regardless of their antiphospholipid reactivities. Macrophages from patients with significant titers of anti- PS IgG antibodies were not activated by the presence of apoptotic CEM lymphoblasts or by purified anti-PS IgG from the same patients. By contrast, these antibodies greatly improved the effector functions of autologous macrophages in antibody-dependent cellular cytotoxicity (ADCC) assays using 51Cr-labeled CEM cells, whereas polyspecific IgG were unable to induce an equivalent cytotoxicity in all instances. An increasing effect on ADCC was also observed in tests using macrophages from healthy controls to CEM coated with anti-PS IgG. These results support a potential correlation of anti-PS specificity with T-cell apoptosis in HIV-1 infection. Because PS is exteriorized by apoptotic lymphocytes, its persistence may stimulate antibodies which cooperate with macrophages in the clearance of dead cells by an enhanced ADCC mechanism. This interpretation could explain the absence of thrombophilia in HIV-1+ patients with serum elevations of antiphospholipid reactivities.

scholarly journals Cross-Reactive Human Immunodeficiency Virus Type 1-Neutralizing Human Monoclonal Antibody That Recognizes a Novel Conformational Epitope on gp41 and Lacks Reactivity against Self-Antigens

2008 ◽  
Vol 82 (14) ◽  
pp. 6869-6879 ◽  
Author(s):  
Mei-Yun Zhang ◽  
Bang K. Vu ◽  
Anil Choudhary ◽  
Hong Lu ◽  
Michael Humbert ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Human Immunodeficiency Virus ◽  
Conformational Epitope ◽  
Heptad Repeat ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Antibody ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Self Antigens

ABSTRACT Broadly cross-reactive human immunodeficiency virus (HIV)-neutralizing antibodies are infrequently elicited in infected humans. The two best-characterized gp41-specific cross-reactive neutralizing human monoclonal antibodies, 4E10 and 2F5, target linear epitopes in the membrane-proximal external region (MPER) and bind to cardiolipin and several other autoantigens. It has been hypothesized that, because of such reactivity to self-antigens, elicitation of 2F5 and 4E10 and similar antibodies by vaccine immunogens based on the MPER could be affected by tolerance mechanisms. Here, we report the identification and characterization of a novel anti-gp41 monoclonal antibody, designated m44, which neutralized most of the 22 HIV type 1 (HIV-1) primary isolates from different clades tested in assays based on infection of peripheral blood mononuclear cells by replication-competent virus but did not bind to cardiolipin and phosphatidylserine in an enzyme-linked immunosorbent assay and a Biacore assay nor to any protein or DNA autoantigens tested in Luminex assays. m44 bound to membrane-associated HIV-1 envelope glycoproteins (Envs), to recombinant Envs lacking the transmembrane domain and cytoplasmic tail (gp140s), and to gp41 structures containing five-helix bundles and six-helix bundles, but not to N-heptad repeat trimers, suggesting that the C-heptad repeat is involved in m44 binding. In contrast to 2F5, 4E10, and Z13, m44 did not bind to any significant degree to denatured gp140 and linear peptides derived from gp41, suggesting a conformational nature of the epitope. This is the first report of a gp41-specific cross-reactive HIV-1-neutralizing human antibody that does not have detectable reactivity to autoantigens. Its novel conserved conformational epitope on gp41 could be helpful in the design of vaccine immunogens and as a target for therapeutics.

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