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1.
Catherine Spenlehauer Sentob Saragosti Herv J. A. Fleury Andr Kirn Anne-Marie Aubertin Christiane Moog 《Journal of virology》1998,72(12):9855-9864
Previous studies characterized the third variable (V3) loop of the envelope gp120 as the principal neutralizing determinant for laboratory T-cell-line-adapted (TCLA) strains of human immunodeficiency virus type 1 (HIV-1). However, primary viruses isolated from infected individuals are more refractory to neutralization than TCLA strains, suggesting that qualitatively different neutralizing antibodies may be involved. In this study, we investigated whether the V3 loop constitutes a linear target epitope for antibodies neutralizing primary isolates. By using peptides representative of the V3 regions of various primary isolates, an early, relatively specific and persistent antibody response was detected in sera from HIV-infected patients. To assess the relationship between these antibodies and neutralization, the same peptides were used in competition and depletion experiments. Addition of homologous V3 peptides led to a competitive inhibition in the neutralization of the TCLA strain HIVMN/MT-4 but had no effect on the neutralization of the autologous primary isolate. Similarly, the removal of antibodies that bind to linear V3 epitopes resulted in a loss of HIVMN/MT-4 neutralization, whereas no decrease in the autologous neutralization was measured. The different roles of V3-specific antibodies according to the virus considered were thereby brought to light. This confirmed the involvement of V3 antibodies in the neutralization of a TCLA strain but emphasized a more pronounced contribution of either conformational epitopes or epitopes outside the V3 loop as targets for antibodies neutralizing primary HIV-1 isolates. This result underlines the need to focus on new vaccinal immunogens with epitopes able to induce broadly reactive and efficient antibodies that neutralize a wide range of primary HIV-1 isolates. 相似文献
2.
Morris L Chen X Alam M Tomaras G Zhang R Marshall DJ Chen B Parks R Foulger A Jaeger F Donathan M Bilska M Gray ES Abdool Karim SS Kepler TB Whitesides J Montefiori D Moody MA Liao HX Haynes BF 《PloS one》2011,6(9):e23532
Broadly neutralizing antibodies are not commonly produced in HIV-1 infected individuals nor by experimental HIV-1 vaccines. When these antibodies do occur, it is important to be able to isolate and characterize them to provide clues for vaccine design. CAP206 is a South African subtype C HIV-1-infected individual previously shown to have broadly neutralizing plasma antibodies targeting the envelope gp41 distal membrane proximal external region (MPER). We have now used a fluoresceinated peptide tetramer antigen with specific cell sorting to isolate a human neutralizing monoclonal antibody (mAb) against the HIV-1 envelope gp41 MPER. The isolated recombinant mAb, CAP206-CH12, utilized a portion of the distal MPER (HXB2 amino acid residues, 673-680) and neutralized a subset of HIV-1 pseudoviruses sensitive to CAP206 plasma antibodies. Interestingly, this mAb was polyreactive and used the same germ-line variable heavy (V(H)1-69) and variable kappa light chain (V(K)3-20) gene families as the prototype broadly neutralizing anti-MPER mAb, 4E10 (residues 672-680). These data indicate that there are multiple immunogenic targets in the C-terminus of the MPER of HIV-1 gp41 envelope and suggests that gp41 neutralizing epitopes may interact with a restricted set of naive B cells during HIV-1 infection. 相似文献
3.
Examination of sera from human immunodeficiency virus type 1 (HIV-1)-infected individuals for antibodies reactive with peptides corresponding to the principal neutralizing determinant of HIV-1 gp120 and for in vitro neutralizing activity.
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R Q Warren S A Anderson W M Nkya J F Shao C W Hendrix G P Melcher R R Redfield R C Kennedy 《Journal of virology》1992,66(9):5210-5215
Sera from human immunodeficiency virus type 1 (HIV-1)-infected individuals from the United States and Tanzania were examined for antibody reactivity to four synthetic peptides which corresponded to the principal neutralizing determinant from the V3 region of HIV-1 gp120. We observed that the majority of sera from both countries contained antibodies reactive with a V3 peptide whose sequence is based on that of the HIV-1 MN isolate. We were unable to establish a relationship between the presence of V3-reactive antibodies, as measured by enzyme-linked immunosorbent assay and neutralization of homologous HIV-1 isolates, in sera from either the United States or Tanzania. We observed that some sera which contained high antibody titers to the V3 peptides failed to neutralize HIV-1, while others with no antibody reactivity to the panel of V3 peptides exhibited in vitro neutralizing activity. These results suggest that neutralizing epitopes exist outside the V3 loop and that the presence of V3-reactive antibodies in sera does not imply in vitro neutralization of the homologous HIV-1 isolate. In addition, it appears that the V3 loop may consist of both neutralizing and nonneutralizing epitopes. The identification of neutralizing as well as nonneutralizing epitopes will be important for the design of potential HIV-1 vaccines. 相似文献
4.
Discontinuous, conserved neutralization epitopes overlapping the CD4-binding region of human immunodeficiency virus type 1 gp120 envelope glycoprotein. 总被引:45,自引:35,他引:10
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M Thali C Furman D D Ho J Robinson S Tilley A Pinter J Sodroski 《Journal of virology》1992,66(9):5635-5641
Monoclonal antibodies have been isolated from human immunodeficiency virus type 1 (HIV-1)-infected patients that recognize discontinuous epitopes on the gp120 envelope glycoprotein, that block gp120 interaction with the CD4 receptor, and that neutralize a variety of HIV-1 isolates. Using a panel of HIV-1 gp120 mutants, we identified amino acids important for precipitation of the gp120 glycoprotein by three different monoclonal antibodies with these properties. These amino acids are located within seven discontinuous, conserved regions of the gp120 glycoprotein, four of which overlap those regions previously shown to be important for CD4 recognition. The pattern of sensitivity to amino acid change in these seven regions differed for each antibody and also differed from that of the CD4 glycoprotein. These results indicate that the CD4 receptor and this group of broadly neutralizing antibodies recognize distinct but overlapping gp120 determinants. 相似文献
5.
R Stanfield E Cabezas A Satterthwait E Stura A Profy I Wilson 《Structure (London, England : 1993)》1999,7(2):131-142
BACKGROUND: The third hypervariable (V3) loop of HIV-1 gp120 has been termed the principal neutralizing determinant (PND) of the virus and is involved in many aspects of virus infectivity. The V3 loop is required for viral entry into the cell via membrane fusion and is believed to interact with cell surface chemokine receptors on T cells and macrophages. Sequence changes in V3 can affect chemokine receptor usage, and can, therefore, modulate which types of cells are infected. Antibodies raised against peptides with V3 sequences can neutralize laboratory-adapted strains of the virus and inhibit syncytia formation. Fab fragments of these neutralizing antibodies in complex with V3 loop peptides have been studied by X-ray crystallography to determine the conformation of the V3 loop. RESULTS: We have determined three crystal structures of Fab 58.2, a broadly neutralizing antibody, in complex with one linear and two cyclic peptides the amino acid sequence of which comes from the MN isolate of the gp120 V3 loop. Although the peptide conformations are very similar for the linear and cyclic forms, they differ from that seen for the identical peptide bound to a different broadly neutralizing antibody, Fab 59.1, and for a similar peptide bound to the MN-specific Fab 50.1. The conformational difference in the peptide is localized around residues Gly-Pro-Gly-Arg, which are highly conserved in different HIV-1 isolates and are predicted to adopt a type II beta turn. CONCLUSIONS: The V3 loop can adopt at least two different conformations for the highly conserved Gly-Pro-Gly-Arg sequence at the tip of the loop. Thus, the HIV-1 V3 loop has some inherent conformational flexibility that may relate to its biological function. 相似文献
6.
Strain specificity and binding affinity requirements of neutralizing monoclonal antibodies to the C4 domain of gp120 from human immunodeficiency virus type 1. 总被引:7,自引:7,他引:0
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G R Nakamura R Byrn D M Wilkes J A Fox M R Hobbs R Hastings H C Wessling M A Norcross B M Fendly P W Berman 《Journal of virology》1993,67(10):6179-6191
The binding properties of seven CD4-blocking monoclonal antibodies raised against recombinant gp120 of human immunodeficiency virus type 1 strain MN (HIV-1MN) and two CD4-blocking monoclonal antibodies to recombinant envelope glycoproteins gp120 and gp160 of substrain IIIB of HIVLAI were analyzed. With a panel of recombinant gp120s from seven diverse HIV-1 isolates, eight of the nine antibodies were found to be strain specific and one was broadly cross-reactive. Epitope mapping revealed that all nine antibodies bound to epitopes located in the fourth conserved domain (C4) of gp120. Within this region, three distinct epitopes could be identified: two were polymorphic between HIV-1 strains, and one was highly conserved. Studies with synthetic peptides demonstrated that the conserved epitope, recognized by antibody 13H8, was located between residues 431 and 439. Site-directed mutagenesis of gp120 demonstrated that residue 429 and/or 432 was critical for the binding of the seven antibodies to gp120 from HIV-1MN. Similarly, residues 423 and 429 were essential for the binding of monoclonal antibody 5C2 raised against gp120 from HIV-1IIIB. The amino acids located at positions 423 and 429 were found to vary between strains of HIV-1 as well as between molecular clones derived from the MN and LAI isolates of HIV-1. Polymorphism at these positions prevented the binding of virus-neutralizing monoclonal antibodies and raised the possibility that HIV-1 neutralization serotypes may be defined on the basis of C4 domain sequences. Analysis of the binding characteristics of the CD4-blocking antibodies demonstrated that their virus-neutralizing activity was directly proportional to their gp120-binding affinity. These studies account for the strain specificity of antibodies to the C4 domain of gp120 and demonstrate for the first time that antibodies to this region can be as effective as those directed to the principal neutralizing determinant (V3 domain) in neutralizing HIV-1 infectivity. 相似文献
7.
Localization of neutralizing regions of the envelope gene of feline leukemia virus by using anti-synthetic peptide antibodies. 总被引:4,自引:3,他引:1
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J H Elder J S McGee M Munson R A Houghten W Kloetzer J L Bittle C K Grant 《Journal of virology》1987,61(1):8-15
We synthesized 27 synthetic peptides corresponding to approximately 80% of the sequences encoding gp70 and p15E of Gardner-Arnstein feline leukemia virus (FeLV) subtype B. The peptides were conjugated to keyhole limpet hemocyanin and injected into rabbits for preparation of antipeptide antisera. These sera were then tested for their ability to neutralize a broad range of FeLV isolates in vitro. Eight peptides elicited neutralizing responses against subtype B isolates. Five of these peptides corresponded to sequences of gp70 and three to p15E. The ability of these antipeptide antisera to neutralize FeLV subtypes A and C varied. In certain circumstances, failure to neutralize a particular isolate corresponded to sequence changes within the corresponding peptide region. However, four antibodies which preferentially neutralized the subtype B viruses were directed to epitopes in common with Sarma subtype C virus. These results suggest that distal changes in certain subtypes (possibly glycosylation differences) alter the availability of certain epitopes in one virus isolate relative to another. We prepared a "nest" of overlapping peptides corresponding to one of the neutralizing regions of gp70 and performed slot blot analyses with both antipeptide antibodies and a monoclonal antibody which recognized this epitope. We were able to define a five-amino-acid sequence required for reactivity. Comparisons were made between an anti-synthetic peptide antibody and a monoclonal antibody reactive to this epitope for the ability to bind both peptide and virus, as well as to neutralize virus in vitro. Both the anti-synthetic peptide and the monoclonal antibodies bound peptide and virus to high titers. However, the monoclonal antibody had a 4-fold-higher titer against virus and a 10-fold-higher neutralizing titer than did the anti-synthetic peptide antibody. Competition assays were performed with these two antibodies adjusted to equivalent antivirus titers against intact virions affixed to tissue culture plates. The monoclonal antibody had a greater ability to compete for virus binding, which suggested that differences in neutralizing titers may relate to the relative affinities of these antisera for the peptide conformation in the native structure. 相似文献
8.
Dissecting the neutralizing antibody specificities of broadly neutralizing sera from human immunodeficiency virus type 1-infected donors
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Dhillon AK Donners H Pantophlet R Johnson WE Decker JM Shaw GM Lee FH Richman DD Doms RW Vanham G Burton DR 《Journal of virology》2007,81(12):6548-6562
Attempts to elicit broadly neutralizing antibody responses by human immunodeficiency virus type 1 (HIV-1) vaccine antigens have been met with limited success. To better understand the requirements for cross-neutralization of HIV-1, we have characterized the neutralizing antibody specificities present in the sera of three asymptomatic individuals exhibiting broad neutralization. Two individuals were infected with clade B viruses and the third with a clade A virus. The broadly neutralizing activity could be exclusively assigned to the protein A-reactive immunoglobulin G (IgG) fraction of all three donor sera. Neutralization inhibition assays performed with a panel of linear peptides corresponding to the third hypervariable (V3) loop of gp120 failed to inhibit serum neutralization of a panel of HIV-1 viruses. The sera also failed to neutralize chimeric simian immunodeficiency virus (SIV) and HIV-2 viruses displaying highly conserved gp41-neutralizing epitopes, suggesting that antibodies directed against these epitopes likely do not account for the broad neutralizing activity observed. Polyclonal IgG was fractionated on recombinant monomeric clade B gp120, and the neutralization capacities of the gp120-depleted samples were compared to that of the original polyclonal IgG. We found that the gp120-binding antibody population mediated neutralization of some isolates, but not all. Overall, the data suggest that broad neutralization results from more than one specificity in the sera but that the number of these specificities is likely small. The most likely epitope recognized by the monomeric gp120 binding neutralizing fraction is the CD4 binding site, although other epitopes, such as the glycan shield, cannot be excluded. 相似文献
9.
Variant-specific monoclonal and group-specific polyclonal human immunodeficiency virus type 1 neutralizing antibodies raised with synthetic peptides from the gp120 third variable domain. 总被引:6,自引:0,他引:6
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J D Laman M M Schellekens Y H Abacioglu G K Lewis M Tersmette R A Fouchier J P Langedijk E Claassen W J Boersma 《Journal of virology》1992,66(3):1823-1831
The third variable (V3) domain of the human immunodeficiency virus type 1 (HIV-1) external membrane glycoprotein gp120 is of crucial importance in eliciting neutralizing antibodies in infected persons. Polyclonal (PAb) and monoclonal (MAb) antibodies directed against selected epitopes in the V3 domain are valuable tools for analysis of the involvement of such sequences in neutralization and for definition of the relation between amino acid variability and immunological cross-reactions. The aim of this study was to obtain such site-specific antibodies. By using synthetic peptides derived from the V3 domain, a group-specific neutralizing PAb, two high-affinity HIV-1 IIIB neutralizing MAb, and two nonneutralizing MAb were raised. A 15-amino-acid peptide overlapping the tip of the V3 domain of HIV-1 MN was used to produce a rabbit PAb (W0/07). This PAb inhibited syncytium formation induced by HIV-1 IIIB and four field isolates. A similar IIIB-derived peptide was used to generate two murine immunoglobulin G1 (IgG1) MAb (IIIB-V3-13 and IIIB-V3-34). Pepscan analysis mapped the binding site of IIIB-V3-34 to the sequence IRIQRGPGR. The Kds of IIIB-V3-13 and IIIB-V3-34 for gp120 were 6.8 x 10(-11) and 1.6 x 10(-10) M, respectively. These MAb neutralized IIIB but not MN and inhibited syncytium formation induced by IIIB. They are applicable in enzyme-linked immunosorbent assays, immunocytochemistry, and flow cytometry. A peptide covering the left base of the V3 domain was used to generate two murine IgG1 MAb (IIIB-V3-21 and IIIB-V3-26). The binding site of IIIB-V3-21 was mapped to the sequence INCTRPN. These MAb did not neutralize HIV-1 and did not inhibit syncytium formation. This study supports the notion that HIV-1 neutralizing antibodies suitable for multiassay performance can be obtained with synthetic peptides and that high-affinity MAb can be generated. Such site-specific antibodies are useful reagents in the analysis of HIV-1 neutralization. In addition, the cross-neutralization of different viral strains by PAb generated through single-peptide immunization is directly relevant to vaccine development. 相似文献
10.
Background
Novel strategies are needed for the elicitation of broadly neutralizing antibodies to the HIV envelope glycoprotein, gp120. Experimental evidence suggests that combinations of antibodies that are broadly neutralizing in vitro may protect against challenge with HIV in nonhuman primates, and a small number of these antibodies have been selected by repertoire sampling of B cells and by the fractionation of antiserum from some patients with prolonged disease. Yet no additional strategies for identifying conserved epitopes, eliciting antibodies to these epitopes, and determining whether these epitopes are accessible to antibodies have been successful to date. The defining of additional conserved, accessible epitopes against which one can elicit antibodies will increase the probability that some may be the targets of broadly neutralizing antibodies.Methodology/Principal Findings
We postulate that additional cryptic epitopes of gp120 are present, against which neutralizing antibodies might be elicited even though these antibodies are not elicited by gp120, and that many of these epitopes may be accessible to antibodies should they be formed. We demonstrate a strategy for eliciting antibodies in mice against selected cryptic, conformationally dependent conserved epitopes of gp120 by immunizing with multiple identical copies of covalently linked peptides (MCPs). This has been achieved with MCPs representing 3 different domains of gp120. We show that some cryptic epitopes on gp120 are accessible to the elicited antibodies, and some epitopes in the CD4 binding region are not accessible. The antibodies bind to gp120 with relatively high affinity, and bind to oligomeric gp120 on the surface of infected cells.Conclusions/Significance
Immunization with MCPs comprised of selected peptides of HIV gp120 is able to elicit antibodies against conserved, conformationally dependent epitopes of gp120 that are not immunogenic when presented as gp120. Some of these cryptic epitopes are accessible to the elicited antibodies. 相似文献11.
Extensive glycosylation of the envelope spikes of human and simian immunodeficiency virus (HIV and SIV) is an important factor for the resistance of these viruses to neutralization by antibodies. SIVmac239 gp41 has three closely spaced sites for N-linked carbohydrate attachment. Rhesus macaques experimentally infected with mutant versions of SIVmac239 lacking two or three of these carbohydrate sites developed strong serum reactivity against mutated peptide sequences at the site of these glycosylations, as well as high titers of neutralizing activity to the mutant viruses (E. Yuste et al., J. Virol. 82:12472–12486, 2008). However, whether antibodies that recognize these underlying peptides have neutralizing activity has not been directly demonstrated. Here we describe the isolation and characterization of three gp41-specific monoclonal antibodies (4G8, 6G8, and 7D6) from one of these mutant-infected monkeys. All three antibodies reacted with mutant gp41 from viral particles and also with peptides corresponding to mutated sequences. Slight differences in peptide specificities were observed among the three antibodies. Sequence analysis revealed that the heavy chains of all three antibodies were derived from the same germ line heavy-chain segment (IGHV4-59*01), but they all had very different sequences in complementarity-determining region 3. The light chains of all three antibodies were very closely related to one another. All three antibodies had neutralizing activity to mutant viruses deficient in gp41 carbohydrate attachment, but they did not neutralize the parental SIVmac239. These results demonstrate unambiguously that antibodies with specificity for peptide sequences underlying gp41 carbohydrates can effectively neutralize SIV when these carbohydrates are absent. However, the presence of these gp41 carbohydrates effectively shields the virus from antibodies that would otherwise neutralize viral infectivity. 相似文献
12.
Polyvalent human immunodeficiency virus synthetic immunogen comprised of envelope gp120 T helper cell sites and B cell neutralization epitopes 总被引:14,自引:0,他引:14
T J Palker T J Matthews A Langlois M E Tanner M E Martin R M Scearce J E Kim J A Berzofsky D P Bolognesi B F Haynes 《Journal of immunology (Baltimore, Md. : 1950)》1989,142(10):3612-3619
In previous studies, we have used antisera raised to envelope (env)-gene-encoded synthetic peptides to identify a region of (HIV) glycoprotein (gp) 120 env protein designated SP10 that contains a type-specific neutralizing determinant. To develop a polyvalent, synthetic peptide inoculum that can evoke both neutralizing antibodies and T cell proliferative responses to more than one HIV isolate, synthetic peptides containing type-specific neutralizing determinants of gp120 from HIV isolates HTLV-IIIB (IIIB), HTLV-IIIMN (MN) and HTLV-IIIRF (RF) were coupled to a 16 amino acid T cell epitope (T1) of HIV-IIIB gp120 and used to immunize goats. Goat antisera to each T1-SP10 peptide derived from the SP10 region of gp120 of IIIB, MN, and RF neutralized HIV isolates IIIB, MN and RF in a type-specific manner. Moreover, peripheral blood T cells from immunized goats also proliferated in a type-specific manner to peptides derived from gp120 of IIIB, MN, and RF. When combined in a trivalent inoculum, T1-SP10 peptides from HIV-1 isolates IIIB, MN, and RF evoked a high titered neutralizing antibody response to isolates IIIB, MN, and RF in goats and as well induced immune T cells to undergo blast transformation in the presence of peptides derived from gp120 of all three HIV isolates. The T1 portion of the T1-SP10 construct was shown to induce a B cell antibody response against determinants within the T1 peptide in addition to inducing T cell proliferative responses in immune goat T cells. Moreover, the SP10 portion of the T1-SP10 constructs not only induced B cell antibody production but also induced type-specific T cell proliferative responses localized to the C-terminal variable sequences of the SP10 peptides. Finally, the T1-SP10 peptide construct induced memory T cell proliferative responses to native gp120 env protein. Thus, combinations of homologous SP10 region synthetic peptides containing type-specific neutralizing determinants and T cell epitopes of HIV gp120 may be useful in man to elicit high titered neutralizing B cell responses and, as well, T cell responses to more than one HIV isolate. 相似文献
13.
Immunization with a soluble CD4-gp120 complex preferentially induces neutralizing anti-human immunodeficiency virus type 1 antibodies directed to conformation-dependent epitopes of gp120. 总被引:7,自引:6,他引:1
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Preservation of the conformation of recombinant gp120 in an adjuvant, enabling it to elicit conformation-dependent, epitope-specific, broadly neutralizing antibodies, may be critical for the development of any gp120-based human immunodeficiency virus type 1 (HIV-1) vaccine. It was hypothesized that recombinant gp120 complexed with recombinant CD4 could stabilize the conformation-dependent neutralizing epitopes and effectively deliver them to the immune system. Therefore, a soluble CD4-gp120 complex in Syntex adjuvant formulation was tested with mice for its ability to induce neutralizing anti-gp120 antibody responses. Seventeen monoclonal antibodies (MAbs) were generated and characterized. Immunochemical studies, neutralization assays, and mapping studies with gp120 mutants indicated that the 17 MAbs fell into three groups. Four of them were directed to what is probably a conformational epitope involving the C1 domain and did not possess virus-neutralizing activities. Another four MAbs bound to V3 peptide 302-321 and exhibited cross-reactive gp120 binding and relatively weak virus-neutralizing activities. These MAbs were very sensitive to amino acid substitutions, not only in the V3 regions but also in the base of the V1/V2 loop, implying a conformational constraint on the epitope. The last group of nine MAbs recognized conformation-dependent epitopes near the CD4 binding site of gp120 and inhibited the gp120-soluble CD4 interaction. Four of these nine MAbs showed broadly neutralizing activities against multiple laboratory-adapted strains of HIV-1, three of them neutralized only HIVIIIB, and the two lower-affinity MAbs did not neutralize any strain tested. Collectively, the results from this study indicate that immunization with the CD4-gp120 complex can elicit antibodies to conformationally sensitive gp120 epitopes, with some of the antibodies having broadly neutralizing activities. We suggest that immunization with CD4-gp120 complexes may be worth evaluating further for the development of an AIDS vaccine. 相似文献
14.
Riabinina SA Baranova EN Sharipova IN Susekina MI Puzyrev VF Obriadina AP Burkov AN Ulanova TI 《Molekuliarnaia genetika, mikrobiologiia i virusologiia》2007,(3):33-36
The third hypervariable domain V3 of the human immunodeficiency virus type 1 gpl20 envelope glycoprotein contains neutralizing epitopes and plays an important role in the diagnosis of HIV infection . Neutralizing antibodies bind to conserved epitope with sequence GPG of V3 loop. The effect of sequence variation on the antigenic properties of the V3 epitope gp120 was studied using five synthetic peptides. The amino acid sequence of the peptide corresponding to the V3 region gp120 of HIV-1 subtype C showed the highest immunoreactivity. The DNA fragment encoding V3-C region gp120 was synthesized by polymerase chain reaction and cloned into pET41b vector. The recombinant plasmid was expressed in the E. coli cells, and recombinant protein was purified using glutathione-S sepharose affinity chromatography. The serological activity of the recombinant protein was tested using ELISA and compared to activity of similar synthetic peptide. The results of this study showed that most immunoreactive agent was the amino acid sequence of V3 region gp120 of HIV-1 subtype C. The recombinant antigen comprising this sequence was more antigenic than synthetic peptide with the same sequence. The evaluation of this antigen shows that this protein is a good candidate for the immunoassay development. 相似文献
15.
Detailed mapping of the antigenicity of the surface unit glycoprotein of equine infectious anemia virus by using synthetic peptide strategies. 总被引:15,自引:6,他引:9
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We describe here a detailed analysis of the antigenic determinants of the surface unit glycoprotein (gp90) of equine infectious anemia virus (EIAV), using a comprehensive panel of synthetic peptides in enzyme-linked immunosorbent assays with immune serum from naturally and experimentally infected horses and with a panel of gp90-specific neutralizing and nonneutralizing monoclonal antibodies. The results of these studies identify immunoreactive segments throughout the conserved and variable domains of gp90 but localize immunodominant (100% reactivity) determinants to the amino and carboxyl termini of the glycoprotein molecule. Analysis of peptide reactivities with longitudinal serum samples taken from experimentally infected ponies revealed that antibody responses to conserved B-cell determinants appeared earlier and at higher titers than do antibodies specific for determinants contained in the variable domain of gp90. These observations suggest an evolution of antibody responses in EIAV-infected ponies that may correspond to the establishment of immunological control of virus replication and disease routinely observed in EIAV infections. In addition, the mapping of monoclonal antibody epitopes to peptides of 9 to 12 amino acids demonstrated that all of the neutralizing epitopes are located in the variable domain of gp90. The arrangement of neutralizing epitopes and critical structural considerations suggest that EIAV gp90 contains a principal neutralizing domain similar to the V3 loop of human immunodeficiency virus type 1. These antigenic analyses provide an important foundation for further analyzing the protective immune response generated during persistent EIAV infections and also provide potential peptide substrates for diagnostic assays and for vaccine strategies. 相似文献
16.
Induction of antibodies in guinea pigs and rhesus monkeys against the human immunodeficiency virus type 1 envelope: neutralization of nonpathogenic and pathogenic primary isolate simian/human immunodeficiency virus strains 总被引:1,自引:0,他引:1
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Liao HX Etemad-Moghadam B Montefiori DC Sun Y Sodroski J Scearce RM Doms RW Thomasch JR Robinson S Letvin NL Haynes BF 《Journal of virology》2000,74(1):254-263
We have compared the abilities of human immunodeficiency virus type 1 (HIV-1) envelope V3 peptides and recombinant gp120 to induce antibodies that neutralize simian/human immunodeficiency viruses (SHIVs). SHIV-89.6 is a nonpathogenic SHIV that expresses the envelope protein of primary HIV-1 isolate 89.6. SHIV-89.6P, clone KB9, is a pathogenic SHIV variant derived from SHIV-89.6. Infection of rhesus monkeys with these SHIVs rarely induces anti-V3 region antibodies. To determine the availability of the gp120 V3 loop for neutralizing antibody binding on SHIV-89.6 and KB9 virions, we have constructed immunogenic C4-V3 peptides from these SHIVs and induced anti-V3 antibodies in guinea pigs and rhesus monkeys. We found that both SHIV-89.6 and KB9 C4-V3 peptides induced antibodies that neutralized SHIV-89.6 but that only SHIV-KB9 C4-V3 peptide induced antibodies that neutralized SHIV-KB9. Immunoprecipitation assays demonstrated that SHIV-KB9 C4-V3 peptide-induced antibodies had a greater ability to bind SHIV-KB9 envelope proteins than did antibodies raised against SHIV-89.6 C4-V3 peptide. We have used a series of mutant HIV-1 envelope constructs to map the gp120 determinants that affect neutralization by anti-V3 antibodies. The residue change at position 305 of arginine (in SHIV-89.6) to glutamic acid (in SHIV-KB9) played a central role in determining the ability of peptide-induced anti-V3 antiserum to neutralize primary isolate SHIVs. Moreover, residue changes in the SHIV-89.6 V1/V2 loops also played roles in regulating the availability of the V3 neutralizing epitope on SHIV-89.6 and -KB9. Thus, SHIV-89.6 and -KB9 V3 region peptides are capable of inducing neutralizing antibodies against these primary isolate SHIVs, although the pathogenic SHIV-KB9 is less easily neutralized than its nonpathogenic variant SHIV-89.6. In contrast to natural infection with SHIV-89.6, in which few animals make anti-V3 antibodies, C4-V3 peptides frequently induced anti-V3 antibodies that neutralized primary isolate SHIV strains. 相似文献
17.
Characterization of a discontinuous human immunodeficiency virus type 1 gp120 epitope recognized by a broadly reactive neutralizing human monoclonal antibody. 总被引:53,自引:42,他引:11
M Thali U Olshevsky C Furman D Gabuzda M Posner J Sodroski 《Journal of virology》1991,65(11):6188-6193
While one hypervariable, linear neutralizing determinant on the human immunodeficiency virus type 1 (HIV-1) gp120 envelope glycoprotein has been well characterized, little is known about the conserved, discontinuous gp120 epitopes recognized by neutralizing antibodies in infected individuals. Here, the epitope recognized by a broadly reactive neutralizing monoclonal antibody (F105) derived from an HIV-1-infected patient was characterized by examining the effects of changes in conserved gp120 amino acids on antibody reactivity. The F105 epitope was disrupted by changes in gp120 amino acids 256 and 257, 368 to 370, 421, and 470 to 484, which is consistent with the discontinuous nature of the epitope. Three of these regions are proximal to those previously shown to be important for CD4 binding, which is consistent with the ability of the F105 antibody to block gp120-CD4 interaction. Since F105 recognition was more sensitive to amino acid changes in each of the four identified gp120 regions than was envelope glycoprotein function, replication-competent mutant viruses that escaped neutralization by the F105 antibody were identified. These studies identify a conserved, functional HIV-1 gp120 epitope that is immunogenic in man and may serve as a target for therapeutic or prophylactic intervention. 相似文献
18.
Anton N. Chikaev Anastasiya Yu. Bakulina Ryan C. Burdick Larisa I. Karpenko Vinay K. Pathak Alexander A. Ilyichev 《PloS one》2015,10(3)
The ability to induce anti-HIV-1 antibodies that can neutralize a broad spectrum of viral isolates from different subtypes seems to be a key requirement for development of an effective HIV-1 vaccine. The epitopes recognized by the most potent broadly neutralizing antibodies that have been characterized are largely discontinuous. Mimetics of such conformational epitopes could be potentially used as components of a synthetic immunogen that can elicit neutralizing antibodies. Here we used phage display technology to identify peptide motifs that mimic the epitope recognized by monoclonal antibody VRC01, which is able to neutralize up to 91% of circulating primary isolates. Three rounds of biopanning were performed against 2 different phage peptide libraries for this purpose. The binding specificity of selected phage clones to monoclonal antibody VRC01 was estimated using dot blot analysis. The putative peptide mimics exposed on the surface of selected phages were analyzed for conformational and linear homology to the surface of HIV-1 gp120 fragment using computational analysis. Corresponding peptides were synthesized and checked for their ability to interfere with neutralization activity of VRC01 in a competitive inhibition assay. One of the most common peptides selected from 12-mer phage library was found to partially mimic a CD4-binding loop fragment, whereas none of the circular C7C-mer peptides was able to mimic any HIV-1 domains. However, peptides identified from both the 12-mer and C7C-mer peptide libraries showed rescue of HIV-1 infectivity in the competitive inhibition assay. The identification of epitope mimics may lead to novel immunogens capable of inducing broadly reactive neutralizing antibodies. 相似文献
19.
Induction of antibody responses that neutralize human T-cell leukemia virus type I infection in vitro and in vivo by peptide immunization. 总被引:4,自引:3,他引:1
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Y Tanaka R Tanaka E Terada Y Koyanagi N Miyano-Kurosaki N Yamamoto E Baba M Nakamura H Shida 《Journal of virology》1994,68(10):6323-6331
In order to define neutralization regions on the envelope antigen of human T-cell leukemia virus type I (HTLV-I), we have generated a number of new anti-envelope gp46 monoclonal antibodies from rats and mice. Epitopes recognized by new monoclonal antibodies which could neutralize HTLV-I in syncytium and transformation inhibition assays were localized to sequences in gp46 from amino acids 186 to 193, 190 to 195, 191 to 195, 191 to 196, and 194 to 199. Ovalbumin-conjugated synthetic gp46 peptides containing these neutralization epitopes, pep190-199 (a synthetic gp46 peptide containing amino acids 190 to 199) and pep180-204, but not pep185-194 or pep194-203, could give rise to HTLV-I-neutralizing antibody responses in rabbits. These immune or nonimmune rabbits were then challenged with HTLV-I by intravenous inoculation with 5 x 10(7) live HTLV-I-producing ILT-8M2 cells. By a PCR assay, it was revealed that HTLV-I provirus was detected in peripheral blood lymphocytes from nonimmune and pep288-312-immunized rabbits, whereas the provirus was not detected in peripheral blood lymphocytes from pep190-199- and pep180-204-immunized rabbits over an extended period. These results suggest that the induction of anti-gp46 neutralizing antibody responses by immunization with synthetic peptides has the potential to protect animals against HTLV-I infection in vivo. 相似文献
20.
Mapping of homologous, amino-terminal neutralizing regions of human T-cell lymphotropic virus type I and II gp46 envelope glycoproteins. 总被引:6,自引:6,他引:0
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T J Palker E R Riggs D E Spragion A J Muir R M Scearce R R Randall M W McAdams A McKnight P R Clapham R A Weiss et al. 《Journal of virology》1992,66(10):5879-5889
Twelve synthetic peptides containing hydrophilic amino acid sequences of human T-cell lymphotropic virus type I (HTLV-I) envelope glycoprotein were coupled to tetanus toxoid and used to raise epitope-specific antisera in goats and rabbits. Low neutralizing antibody titers (1:10 to 1:20) raised in rabbits to peptides SP-2 (envelope amino acids [aa] 86 to 107), SP-3 (aa 176 to 189), and SP-4A (aa 190 to 209) as well as to combined peptide SP-3/4A (aa 176 to 209) were detected in the vesicular stomatitis virus-HTLV-I pseudotype assay. Higher-titered neutralizing antibody responses to HTLV-I (1:10 to 1:640) were detected with pseudotype and syncytium inhibition assays in four goats immunized with a combined inoculum containing peptides SP-2, SP-3, and SP-4A linked to tetanus toxoid. These neutralizing anti-HTLV-I antibodies were type specific in that they did not inhibit HTLV-II syncytium formation. Neutralizing antibodies in sera from three goats could be absorbed with peptide SP-2 (aa 86 to 107) as well as truncated peptides containing envelope aa 90 to 98, but not with equimolar amounts of peptides lacking envelope aa 90 to 98. To map critical amino acids that contributed to HTLV-I neutralization within aa 88 to 98, peptides in which each amino acid was sequentially replaced by alanine were synthesized. The resulting 11 synthetic peptides with single alanine substitutions were then used to absorb three neutralizing goat antipeptide antisera. Both asparagines at positions 93 and 95 were required for adsorption of neutralizing anti-HTLV-I antibodies from all three sera. Peptide DP-90, containing the homologous region of HTLV-II envelope glycoprotein (aa 82 to 97), elicited antipeptide neutralizing antibodies to HTLV-II in goats that were type specific. In further adsorption experiments, it was determined that amino acid differences between homologous HTLV-I and HTLV-II envelope sequences at HTLV-I aa 95 (N to Q) and 97 (G to L) determined the type specificity of these neutralizing sites. Thus, the amino-terminal regions of HTLV-I and -II gp46 contain homologous, linear, neutralizing determinants that are type specific. 相似文献