- Research
- Open Access
Immunogenicity of the outer domain of a HIV-1 clade C gp120
- Hongying Chen1,
- Xiaodong Xu1 and
- Ian M Jones1Email author
https://doi.org/10.1186/1742-4690-4-33
© Chen et al; licensee BioMed Central Ltd. 2007
- Received: 06 April 2007
- Accepted: 17 May 2007
- Published: 17 May 2007
Abstract
Background
The possibility that a sub domain of a C clade HIV-1 gp120 could act as an effective immunogen was investigated. To do this, the outer domain (OD) of gp120CN54 was expressed and characterized in a construct marked by a re-introduced conformational epitope for MAb 2G12. The expressed sequence showed efficient epitope retention on the isolated ODCN54 suggesting authentic folding. To facilitate purification and subsequent immunogenicity ODCN54 was fused to the Fc domain of human IgG1. Mice were immunised with the resulting fusion proteins and also with gp120CN54-Fc and gp120 alone.
Results
Fusion to Fc was found to stimulate antibody titre and Fc tagged ODCN54 was substantially more immunogenic than non-tagged gp120. Immunogenicity appeared the result of Fc facilitated antigen processing as immunisation with an Fc domain mutant that reduced binding to the FcR lead to a reduction in antibody titre when compared to the parental sequence. The breadth of the antibody response was assessed by serum reaction with five overlapping fragments of gp120CN54 expressed as GST fusion proteins in bacteria. A predominant anti-inner domain and anti-V3C3 response was observed following immunisation with gp120CN54-Fc and an anti-V3C3 response to the ODCN54-Fc fusion.
Conclusion
The outer domain of gp120CN54 is correctly folded following expression as a C terminal fusion protein. Immunogenicity is substantial when targeted to antigen presenting cells but shows V3 dominance in the polyvalent response. The gp120 outer domain has potential as a candidate vaccine component.
Keywords
- Outer Domain
- 2G12 Binding
- Unliganded Gp120
- V3C3 Region
- 2G12 Epitope
Introduction
The need for a form of HIV-1 envelope protein capable of eliciting a broadly neutralising antibody (NAb) response as part of an HIV vaccine has been widely discussed [1–3]. It is generally agreed that the lack of NAb is a consequence of a number of evasion mechanisms evolved by the virus to maintain immunological silence. Examples include glycan shrouding [4] and envelope structural heterogeneity [5, 6]. Envelope structural heterogeneity involves primarily the outer envelope protein gp120. The monomeric molecule is flexible, with comparisons between the crystal structures of liganded and unliganded gp120 showing significant local change [7, 8]. Of the three defined structural domains, the inner domain, bridging sheet and the outer domain (OD), both the inner domain and bridging sheet rearrange substantially upon CD4 binding [8]. The inner domain and bridging sheet are also the source of heterogeneity within monomeric gp120 in solution [6] and have sufficient flexibility to allow structural complementation between adjacent molecules [9, 10]. There is additional heterogeneity in the envelope molecules on the virion surface where gp120 and transmembrane gp41 make up the virion spike which appears patchily distributed and in a variety of conformations [5, 11, 12]. Strategies for improving gp120 immunogenicity have been reviewed recently [13]. One approach has examined the gp120 OD in isolation as, by contrast with the complete molecule, it is structurally stable [14]. The OD is heavily glycosylated and relatively immunologically silent in infected individuals but its potential to act as a generator of NAb is demonstrated by the fact that the epitope for a broad ranging, neutralizing human monoclonal antibody, 2G12, maps to it [15, 16] as do a number of lectins which potently neutralize virus infectivity in vitro [17, 18]. MAb 2G12 is unique in that recognition of its epitope, a high-mannose carbohydrate cluster on gp120, is achieved through a dimeric antibody structure in which there is VH exchange between adjacent immunoglobulin molecules [19]. The paucity of this form of Ig structure in total IgG [20] suggests that a similar specificity may be difficult to generate following immunisation but, as the juxtaposition of key N-linked glycans is essential for MAb binding, the 2G12 epitope does provide a sensitive measure of OD conformation. Yang et al., showed that the OD (residues 252RPVVST.....DNWRS482) of gp120 from B clade virus YU2, which they termed OD1, retained 2G12 binding despite being poorly detected by the majority of HIV positive sera [14, 21]. Interest in the domain lies in the fact that 2G12 does not directly compete for the primary receptor binding site on gp120 [19] but appears to impair secondary receptor binding [22]. In addition, gp120-2G12 complexes exhibited reduced binding to DC-SIGN, consistent with antibody capping of at least some of the mannose moieties that would otherwise bind the lectin [22]. Such in vitro inhibition of gp120 receptor binding is of consequence in vivo as 2G12, in combination with other neutralizing MAbs, such as b12 (against the CD4 binding site [23]) and 2F5 (against gp41), provides protection against HIV-1 challenge in animal models [24–26]. M-type HIV-1 includes 9 clades [27–29] and vaccine candidates designed for beneficial antibody induction should provide immunity against all clades if they are to be effective [3]. It has been noted that C clade envelopes are significantly different to those of B clade isolates, especially early in infection [30, 31] and, more generally, that subtype C candidate vaccine development has been reportedly more problematic than clade B focused strategies [32]. HIV-1 C clade isolates have frequently lost the carbohydrate sites required by 2G12 and so rarely present the epitope [33]. It was unclear therefore if data obtained with the OD of the B clade gp120YU2 [14] would be repeated using the OD of a C-clade isolate nor yet how a meaningful conformation of the latter OD could be confirmed in order to test such a possibility. Here, using the gp120 sequence of HIV-1CN54, a B/C clade recombinant originally isolated in China [34, 35], we examine the C clade gp120 OD using a re-engineered 2G12 epitope to provide a measure of conformational relevance before use as an immunogen. Subsequently, Fc tagged OD is used as an immunogen in a comparative study with gp120-Fc and untagged gp120.
Results and discussion
Characterisation of the 2G12 epitope reconstructed in clade C gp120
Characterisation of HIV-1 gp120CN54 (V295N+A394T) expressed as a transmembrane bound protein in both Spodoptera frugiperda (Sf9) and Mimic cells (Invitrogen). A. Western blot of cell extracts (tracks 1 and 2) and supernatant (tracks 3 and 4) showing the increase in size associated with gp120 with modified glycans. Track 1 and 3 – Sf9 cells, Track 2 and 4 – Mimic cells. Note the presence of the unglycosylated apoprotein (arrowed) and glycosylation intermediates in the cell extracts which are absent in the supernatant. The gp120 present in the supernatant is due to pickup by budding baculovirus particles (see [53]). Markers to the left are in kilodaltons. B. Flow cytometry of permeablized or non-permeablized Sf9 and Mimic cells following staining with 2G12. Cells were permeablized by incubation in 0.1% Triton X-100 followed by fixing in 3% paraformaldehyde. Control cell profiles are shaded, infected cell profiles are unshaded.
The 2G12 epitope is maintained on the isolated outer domain
The juxtaposition of mannose residues on the termini of key gp120 glycans creates the 2G12 epitope which is therefore conformational [15, 16, 21, 33]. Yang et al., described maintenance of the 2G12 epitope on the OD of HIV-1YU2 [14] which also defined the OD as conformationally relevant. To assess if this was also true of the re-engineered 2G12 epitope on gp120CN54+, the OD of gp120CN54 (residues251IKPV...NWRS481) was amplified from a clone encoding gp120CN54+, cloned (additional file 1B)
Purification of gp120CN54+-Fc and ODCN54+-Fc and reaction with MAbs b12 and 2G12. A. Protein present in the supernatant of recombinant baculovirus infected insect cells was purified by lectin and protein A chromatography and analysed by SDS-PAGE (tracks 1 and 2) and western blot (tracks 3 and 4) using an anti-human Ig conjugate. Tracks 1 and 3 – gp120CN54+-Fc;Tracks 2 and 4 – ODCN54+-Fc. Protein size markers indicated to the left are in kilodaltons (kDa). B. Purified protein was coated onto plastic at 10 μg/ml and used to assess the binding of monoclonal antibodies b12 or 2G12 as shown. The samples were (▲) – gp120CN54+-Fc; (■) – ODCN54+-Fc; (◆) – gp120CN54+. Binding was detected by an anti-human Ig light chain conjugate.
Immunogenicity of ODCN54+-Fc, gp120CN54+ and gp120CN54+-Fc
Analysis of serum response to various CN54 gp120s by ELISA and Western blot. A. Pooled mouse sera were assayed on highly purified CN54 gp120 in a reciprocal dilution series starting at 1:100 and binding detected with an anti-mouse conjugate. The immunogens were: (▲) – gp120CN54+-Fc;(■) – ODCN54+-Fc;(◆) – gp120CN54+. B. Western blot analysis of each serum on gp120CN54+ (track 1) ODCN54+-Fc (track 2) and gp120Bal (track3 – to assess cross clade reactivity). The panels were blotted with the sera raised to the proteins shown and were used at the dilutions indicated. Anti-Fc reactivity in the sera was blocked by pre-incubation with excess HCV E2-Fc (not described).
Analysis of serum response to GST-gp120CN54 fragments by Western blot. Expression of each GST fusion protein was induced for 3 hrs and the equivalent of 50 μl of the culture fractionated on 10% SDS-PAGE before the transfer. Lanes 1–5 are the GST-gp120CN54 fusion proteins 1 to 5 with the origin of the gp120CN54 fragments as shown in additional file 2. Lane 6 is GST only (western blot reaction at 26 kDa and 54 kDa in panel A are monomer and dimer of GST respectively). The GST serum (Sigma) was used at 1:5000, serum dilutions for blots B-D were as described for figure 3.
Purification of ODCN54+-Fc, ODCN54+-Fc(VA) and ODCN54+-His. Protein present in the supernatant of recombinant baculovirus infected insect cells was purified by a combination of lectin and protein A chromatography and analysed by 10% SDS-PAGE. Track M – markers with molecular weights as shown; track 1 – ODCN54+-Fc; track 2 – ODCN54+-Fc(VA) and track 3 – ODCN54+-His. Note the slightly slower migration of the VA mutant as a result of the hydrophobicity change in the loss of two leucines within the Fc coding region. The smear of ODCN54+-His was typical and reflects the high level of glycosylation on a fairly small protein fragment. The yield for all constructs was ~1 mg/L of culture.
Analysis of serum responses to gp120CN54 (A) and ODCN54 (B) assessed by ELISA following immunization with the various ODCN54constructs described. Protein immunogens, as shown in figure 5 plus the complex between ODCN54+-Fc and GG7, were used to immunise groups of mice as before and terminal serum titres obtained following incubation on immobilised gp120CN54 and ODCN54 followed by an anti mouse conjugate. The sera were generated to: ODCN54+-Fc (◆); ODCN54+-Fc + GG7 (■); ODCN54+-Fc(VA) (▲) and ODCN54+-His (X).
This data confirms that the outer domain of gp120 in isolation can be immunogenic despite not normally generating significant responses in HIV-1 infected individuals. Similarly, the finding that the majority of the response was to polypeptide not carbohydrate despite the hyperglycosylation of the OD [14] is supported by the reaction of the sera generated with GST-gp120 fusion proteins expressed in bacteria. However, our data do not confirm the conclusion that the OD of HIV-1YU2, upon immunisation, resulted in few antibodies to the V3 loop [14]. That conclusion was inferred from serum depletion experiments and no direct epitope mapping was done whereas our use of overlapping fragments of gp120CN54 as antigen suggests that, for the ODCN54+, induction of a strong response to the V3C3 region of gp120 took place. The pattern of reactivity was the same irrespective of the titre of the serum obtained suggesting that this region is a prominent feature of the CN54 sequence.
The finding that OD-Fc fusions are improved immunogens is consistent with a mechanism of enhanced immunogenicity through engagement of the FcR on antigen presenting cells. The reduction in titre following immunisation with ODCN54+-Fc(VA) supports this conclusion as does the lack of enhancement by further oligomerization through GG7 cross linking. Although reduced, the serum titre in response to ODCN54+-Fc (VA) was still far higher than ODCN54+-His alone probably as a result of a residual level of FcR binding. Within the context of IgG1 the VA mutation leads to a 10–100 fold reduction in FcR binding [48] effectively meaning that 1–10% of the ODCN54+-Fc(VA) immunogen was capable of generating a serum response which was observed to be ~50% of the level obtained with the standard dose of non mutated ODCN54+-Fc. This suggests that gp120-Fc fusions would be immunogenic at significantly lower doses than those used here (10 μg), an important consideration in vaccine design.
The response to gp120CN54+-Fc included reaction with all 5 fragments probed but a predominant response against the N terminus (the C1 domain) and a fragment encoding the V3 loop. Reaction with the C1 domain is consistent with the flexible gp120 inner domain which stimulates antibodies that are non-neutralising [23]. The OD of the C clade gp120CN54+ presented a 2G12 epitope that bound MAb as well as the full length protein (or equivalent amounts of a B clade gp120 – not shown). As the spatial juxtaposition of glycans on residues 295 and 392 is crucial for MAb binding we infer from this that the OD of gp120CN54+ may be structurally very similar to that of the solved B clade structures [7, 8, 23]. Novel mannosylated compounds able to bind 2G12 have been suggested as synthetic immunogens that may be able to elicit 2G12 like antibody responses [50, 51]. A framework based on the natural target, the gp120 OD domain, would be preferable if immunogenicity could be enhanced and the Fc fusion reported here is a simple method of achieving this with the aim of generating further antibodies with 2G12 like properties. In addition, reaction with the V3C3 region was substantial suggesting that substitution of the V3 loop within ODCN54 with other conserved neutralising epitopes, such as those from gp41, may allow their prominent exposure for the generation of broadly cross clade neutralising Abs as has been attempted recently by grafting into the V1/V2 region of gp120 [52].
Conclusion
The 2G12 epitope is faithfully reconstructed in a clade C gp120 backbone with 2 glycosylation site additions. The epitope is retained on the C clade outer domain where it acts as a marker of outer domain conformation. A C-terminal Fc tag provided for enhanced immunogenicity in the absence of any other adjuvants for full length C clade gp120 and for the normally immunologically silent outer domain. The serum response to the outer domain indicated a prominent V3C3 presentation for the clade C molecule in contrast to what has been observed for the clade B outer domain. Conformationally defined gp120 fragments as immune complexes may have potential as part of a vaccine design strategy targeting humoral immunity.
Methods
Cells and manipulations
E. coli Top10 was used for the propagation of plasmids and all cloning. Expression using recombinant baculoviruses used Spodoptera frugiperda (Sf9) insect cells unless otherwise stated. Sf9 cells were cultured in SF900-II (Life Sciences) at 28°C. A description of the cloning and mutagenesis steps is shown in additional file 1.
Recombinant baculovirus infections
Infections for virus growth were done at an MOI of 0.1 and for protein expression, an MOI of 3. Virus growth was typically for 4 days or until there was considerable cytopathic effect. Sf9 cells infected for protein expression were harvested 72 hours post infection and the glycosylated protein present in the supernatant purified as described.
ELISA
Microtitre plates (Thermo Labsystems) were coated with purified proteins previously normalised, blocked with PBS, 5% dried milk powder and used immediately. Primary antibodies were incubated with antigen for 60 min at room temperature. Unbound antibody was removed by washing five times with PBS containing 0.05% v/v Tween-20 and the plate was incubated with HRP-conjugated anti-mouse antibody (1:1000, Chemicon) for one hour at room temperature. The plate was washed extensively and incubated with 3,3',5,5'-Tetramethylbenzidine (TMB) chromagenic substrate (Europa Bioproducts). The reaction was stopped by addition of an equal volume of 0.5 M HCl and the absorbance was read at 410 nm.
Western blotting
Protein samples were separated on pre-cast 10% Tris.HCl SDS-polyacrylamide gels (BioRad) and transferred to Immobilion-P membranes (Millipore) using a semi-dry blotter. Filters were blocked for one hour at room temperature using TBS containing 0.1% v/v Tween-20 (TBS-T), 5% w/v milk powder. Primary antibody was used at a dilution of 1:500 in PBS-T, 5% w/v milk powder unless otherwise stated. Following several washes with TBS-T the membranes were incubated for 1 hour with HRP-conjugated anti-mouse antibody (Chemicon) and the bound antibodies detected by BM chemiluminescence (Roche).
Declarations
Acknowledgements
We thank Herman Katinger and Dennis Burton for the original supplies of 2G12 and b12 respectively and the UK Centre for AIDS Reagents (EU Programme EVA/AVIP) for other HIV reagents. We thank Pam Rummings and Trevor Jenkinson for help with the immunisations. Funding was from the UK Medical Research Council (MRC) and the FP6 microbicide programme of the European Union (EMPRO).
Authors’ Affiliations
References
- Burton DR, Desrosiers RC, Doms RW, Koff WC, Kwong PD, Moore JP, Nabel GJ, Sodroski J, Wilson IA, Wyatt RT: HIV vaccine design and the neutralizing antibody problem. Nat Immunol. 2004, 5 (3): 233-236. 10.1038/ni0304-233.View ArticlePubMedGoogle Scholar
- Burton DR, Stanfield RL, Wilson IA: Antibody vs. HIV in a clash of evolutionary titans. Proc Natl Acad Sci U S A. 2005, 102 (42): 14943-14948. 10.1073/pnas.0505126102.PubMed CentralView ArticlePubMedGoogle Scholar
- Burton DR, Moore JP: Why do we not have an HIV vaccine and how can we make one?. Nat Med. 1998, 4 (5 Suppl): 495-498. 10.1038/nm0598supp-495.View ArticlePubMedGoogle Scholar
- Wei X, Decker JM, Wang S, Hui H, Kappes JC, Wu X, Salazar-Gonzalez JF, Salazar MG, Kilby JM, Saag MS, Komarova NL, Nowak MA, Hahn BH, Kwong PD, Shaw GM: Antibody neutralization and escape by HIV-1. Nature. 2003, 422 (6929): 307-312. 10.1038/nature01470.View ArticlePubMedGoogle Scholar
- Moore PL, Crooks ET, Porter L, Zhu P, Cayanan CS, Grise H, Corcoran P, Zwick MB, Franti M, Morris L, Roux KH, Burton DR, Binley JM: Nature of nonfunctional envelope proteins on the surface of human immunodeficiency virus type 1. J Virol. 2006, 80 (5): 2515-2528. 10.1128/JVI.80.5.2515-2528.2006.PubMed CentralView ArticlePubMedGoogle Scholar
- Yuan W, Bazick J, Sodroski J: Characterization of the multiple conformational States of free monomeric and trimeric human immunodeficiency virus envelope glycoproteins after fixation by cross-linker. J Virol. 2006, 80 (14): 6725-6737. 10.1128/JVI.00118-06.PubMed CentralView ArticlePubMedGoogle Scholar
- Kwong PD, Wyatt R, Robinson J, Sweet RW, Sodroski J, Hendrickson WA: Structure of an HIV gp120 envelope glycoprotein in complex with the CD4 receptor and a neutralizing human antibody. Nature. 1998, 393 (6686): 648-659. 10.1038/31405.View ArticlePubMedGoogle Scholar
- Chen B, Vogan EM, Gong H, Skehel JJ, Wiley DC, Harrison SC: Structure of an unliganded simian immunodeficiency virus gp120 core. Nature. 2005, 433 (7028): 834-841. 10.1038/nature03327.View ArticlePubMedGoogle Scholar
- Pollard SR, Rosa MD, Rosa JJ, Wiley DC: Truncated variants of gp120 bind CD4 with high affinity and suggest a minimum CD4 binding region. Embo J. 1992, 11 (2): 585-591.PubMed CentralPubMedGoogle Scholar
- Morikawa Y, Moore JP, Fenouillet E, Jones IM: Complementation of human immunodeficiency virus glycoprotein mutations in trans. J Gen Virol. 1992, 73 ( Pt 8): 1907-1913.View ArticleGoogle Scholar
- Zhu P, Liu J, Bess J, Chertova E, Lifson JD, Grise H, Ofek GA, Taylor KA, Roux KH: Distribution and three-dimensional structure of AIDS virus envelope spikes. Nature. 2006, 441 (7095): 847-852. 10.1038/nature04817.View ArticlePubMedGoogle Scholar
- Zhu P, Chertova E, Bess J, Lifson JD, Arthur LO, Liu J, Taylor KA, Roux KH: Electron tomography analysis of envelope glycoprotein trimers on HIV and simian immunodeficiency virus virions. Proc Natl Acad Sci U S A. 2003, 100 (26): 15812-15817. 10.1073/pnas.2634931100.PubMed CentralView ArticlePubMedGoogle Scholar
- Pantophlet R, Burton DR: GP120: target for neutralizing HIV-1 antibodies. Annu Rev Immunol. 2006, 24: 739-769. 10.1146/annurev.immunol.24.021605.090557.View ArticlePubMedGoogle Scholar
- Yang X, Tomov V, Kurteva S, Wang L, Ren X, Gorny MK, Zolla-Pazner S, Sodroski J: Characterization of the outer domain of the gp120 glycoprotein from human immunodeficiency virus type 1. J Virol. 2004, 78 (23): 12975-12986. 10.1128/JVI.78.23.12975-12986.2004.PubMed CentralView ArticlePubMedGoogle Scholar
- Sanders RW, Venturi M, Schiffner L, Kalyanaraman R, Katinger H, Lloyd KO, Kwong PD, Moore JP: The mannose-dependent epitope for neutralizing antibody 2G12 on human immunodeficiency virus type 1 glycoprotein gp120. J Virol. 2002, 76 (14): 7293-7305. 10.1128/JVI.76.14.7293-7305.2002.PubMed CentralView ArticlePubMedGoogle Scholar
- Scanlan CN, Pantophlet R, Wormald MR, Ollmann Saphire E, Stanfield R, Wilson IA, Katinger H, Dwek RA, Rudd PM, Burton DR: The broadly neutralizing anti-human immunodeficiency virus type 1 antibody 2G12 recognizes a cluster of alpha1-->2 mannose residues on the outer face of gp120. J Virol. 2002, 76 (14): 7306-7321. 10.1128/JVI.76.14.7306-7321.2002.PubMed CentralView ArticlePubMedGoogle Scholar
- Balzarini J, Van Laethem K, Hatse S, Froeyen M, Van Damme E, Bolmstedt A, Peumans W, De Clercq E, Schols D: Marked depletion of glycosylation sites in HIV-1 gp120 under selection pressure by the mannose-specific plant lectins of Hippeastrum hybrid and Galanthus nivalis. Mol Pharmacol. 2005, 67 (5): 1556-1565. 10.1124/mol.104.005082.View ArticlePubMedGoogle Scholar
- Balzarini J, Van Laethem K, Hatse S, Vermeire K, De Clercq E, Peumans W, Van Damme E, Vandamme AM, Bolmstedt A, Schols D: Profile of resistance of human immunodeficiency virus to mannose-specific plant lectins. J Virol. 2004, 78 (19): 10617-10627. 10.1128/JVI.78.19.10617-10627.2004.PubMed CentralView ArticlePubMedGoogle Scholar
- Calarese DA, Scanlan CN, Zwick MB, Deechongkit S, Mimura Y, Kunert R, Zhu P, Wormald MR, Stanfield RL, Roux KH, Kelly JW, Rudd PM, Dwek RA, Katinger H, Burton DR, Wilson IA: Antibody domain exchange is an immunological solution to carbohydrate cluster recognition. Science. 2003, 300 (5628): 2065-2071. 10.1126/science.1083182.View ArticlePubMedGoogle Scholar
- Roux KH, Zhu P, Seavy M, Katinger H, Kunert R, Seamon V: Electron microscopic and immunochemical analysis of the broadly neutralizing HIV-1-specific, anti-carbohydrate antibody, 2G12. Mol Immunol. 2004, 41 (10): 1001-1011.View ArticlePubMedGoogle Scholar
- Trkola A, Purtscher M, Muster T, Ballaun C, Buchacher A, Sullivan N, Srinivasan K, Sodroski J, Moore JP, Katinger H: Human monoclonal antibody 2G12 defines a distinctive neutralization epitope on the gp120 glycoprotein of human immunodeficiency virus type 1. J Virol. 1996, 70 (2): 1100-1108.PubMed CentralPubMedGoogle Scholar
- Binley JM, Ngo-Abdalla S, Moore P, Bobardt M, Chatterji U, Gallay P, Burton DR, Wilson IA, Elder JH, de Parseval A: Inhibition of HIV Env binding to cellular receptors by monoclonal antibody 2G12 as probed by Fc-tagged gp120. Retrovirology. 2006, 3 (1): 39-10.1186/1742-4690-3-39.PubMed CentralView ArticlePubMedGoogle Scholar
- Zhou T, Xu L, Dey B, Hessell AJ, Van Ryk D, Xiang SH, Yang X, Zhang MY, Zwick MB, Arthos J, Burton DR, Dimitrov DS, Sodroski J, Wyatt R, Nabel GJ, Kwong PD: Structural definition of a conserved neutralization epitope on HIV-1 gp120. Nature. 2007, 445 (7129): 732-737. 10.1038/nature05580.PubMed CentralView ArticlePubMedGoogle Scholar
- Gauduin MC, Parren PW, Weir R, Barbas CF, Burton DR, Koup RA: Passive immunization with a human monoclonal antibody protects hu-PBL-SCID mice against challenge by primary isolates of HIV-1. Nat Med. 1997, 3 (12): 1389-1393. 10.1038/nm1297-1389.View ArticlePubMedGoogle Scholar
- Baba TW, Liska V, Hofmann-Lehmann R, Vlasak J, Xu W, Ayehunie S, Cavacini LA, Posner MR, Katinger H, Stiegler G, Bernacky BJ, Rizvi TA, Schmidt R, Hill LR, Keeling ME, Lu Y, Wright JE, Chou TC, Ruprecht RM: Human neutralizing monoclonal antibodies of the IgG1 subtype protect against mucosal simian-human immunodeficiency virus infection. Nat Med. 2000, 6 (2): 200-206. 10.1038/72309.View ArticlePubMedGoogle Scholar
- Kitabwalla M, Ferrantelli F, Wang T, Chalmers A, Katinger H, Stiegler G, Cavacini LA, Chou TC, Ruprecht RM: Primary African HIV clade A and D isolates: effective cross-clade neutralization with a quadruple combination of human monoclonal antibodies raised against clade B. AIDS Res Hum Retroviruses. 2003, 19 (2): 125-131. 10.1089/088922203762688630.View ArticlePubMedGoogle Scholar
- Robertson DL, Anderson JP, Bradac JA, Carr JK, Foley B, Funkhouser RK, Gao F, Hahn BH, Kalish ML, Kuiken C, Learn GH, Leitner T, McCutchan F, Osmanov S, Peeters M, Pieniazek D, Salminen M, Sharp PM, Wolinsky S, Korber B: HIV-1 nomenclature proposal. Science. 2000, 288 (5463): 55-56. 10.1126/science.288.5463.55d.View ArticlePubMedGoogle Scholar
- van der Groen G, Nyambi PN, Beirnaert E, Davis D, Fransen K, Heyndrickx L, Ondoa P, Van der Auwera G, Janssens W: Genetic variation of HIV type 1: relevance of interclade variation to vaccine development. AIDS Res Hum Retroviruses. 1998, 14 Suppl 3: S211-21.PubMedGoogle Scholar
- Stebbing J, Moyle G: The clades of HIV: their origins and clinical significance. AIDS Rev. 2003, 5 (4): 205-213.PubMedGoogle Scholar
- Kothe DL, Li Y, Decker JM, Bibollet-Ruche F, Zammit KP, Salazar MG, Chen Y, Weng Z, Weaver EA, Gao F, Haynes BF, Shaw GM, Korber BT, Hahn BH: Ancestral and consensus envelope immunogens for HIV-1 subtype C. Virology. 2006, 352 (2): 438-449. 10.1016/j.virol.2006.05.011.View ArticlePubMedGoogle Scholar
- Derdeyn CA, Decker JM, Bibollet-Ruche F, Mokili JL, Muldoon M, Denham SA, Heil ML, Kasolo F, Musonda R, Hahn BH, Shaw GM, Korber BT, Allen S, Hunter E: Envelope-constrained neutralization-sensitive HIV-1 after heterosexual transmission. Science. 2004, 303 (5666): 2019-2022. 10.1126/science.1093137.View ArticlePubMedGoogle Scholar
- Nkolola JP, Essex M: Progress towards an HIV-1 subtype C vaccine. Vaccine. 2006, 24 (4): 391-401. 10.1016/j.vaccine.2005.08.007.View ArticlePubMedGoogle Scholar
- Scanlan CN, Pantophlet R, Wormald MR, Saphire EO, Calarese D, Stanfield R, Wilson IA, Katinger H, Dwek RA, Burton DR, Rudd PM: The carbohydrate epitope of the neutralizing anti-HIV-1 antibody 2G12. Adv Exp Med Biol. 2003, 535: 205-218.View ArticlePubMedGoogle Scholar
- Rodenburg CM, Li Y, Trask SA, Chen Y, Decker J, Robertson DL, Kalish ML, Shaw GM, Allen S, Hahn BH, Gao F: Near full-length clones and reference sequences for subtype C isolates of HIV type 1 from three different continents. AIDS Res Hum Retroviruses. 2001, 17 (2): 161-168. 10.1089/08892220150217247.View ArticlePubMedGoogle Scholar
- Su L, Graf M, Zhang Y, von Briesen H, Xing H, Kostler J, Melzl H, Wolf H, Shao Y, Wagner R: Characterization of a virtually full-length human immunodeficiency virus type 1 genome of a prevalent intersubtype (C/B') recombinant strain in China. J Virol. 2000, 74 (23): 11367-11376. 10.1128/JVI.74.23.11367-11376.2000.PubMed CentralView ArticlePubMedGoogle Scholar
- Chen H, Xu X, Bishop A, Jones IM: Reintroduction of the 2G12 epitope in an HIV-1 clade C gp120. Aids. 2005, 19 (8): 833-835. 10.1097/01.aids.0000168980.74713.9e.View ArticlePubMedGoogle Scholar
- Butters TD, Yudkin B, Jacob GS, Jones IM: Structural characterization of the N-linked oligosaccharides derived from HIVgp120 expressed in lepidopteran cells. Glycoconj J. 1998, 15 (1): 83-88. 10.1023/A:1006999718552.View ArticlePubMedGoogle Scholar
- Jarvis DL: Developing baculovirus-insect cell expression systems for humanized recombinant glycoprotein production. Virology. 2003, 310 (1): 1-7. 10.1016/S0042-6822(03)00120-X.PubMed CentralView ArticlePubMedGoogle Scholar
- Hollister JR, Shaper JH, Jarvis DL: Stable expression of mammalian beta 1,4-galactosyltransferase extends the N-glycosylation pathway in insect cells [In Process Citation]. Glycobiology. 1998, 8 (5): 473-480. 10.1093/glycob/8.5.473.View ArticlePubMedGoogle Scholar
- Centre for AIDS Reagents (EU Programme EVA/AVIP) Internet Catalogue. [http://www.nibsc.ac.uk/spotlight/aidsreagent/index.html]
- Zhao Y, Chapman DA, Jones IM: Improving baculovirus recombination. Nucleic Acids Res. 2003, 31 (2): E6-6. 10.1093/nar/gng006.PubMed CentralView ArticlePubMedGoogle Scholar
- Pengelley SC, Chapman DC, Mark Abbott W, Lin HH, Huang W, Dalton K, Jones IM: A suite of parallel vectors for baculovirus expression. Protein Expr Purif. 2006, 48 (2): 173-181.View ArticlePubMedGoogle Scholar
- Hong PW, Flummerfelt KB, de Parseval A, Gurney K, Elder JH, Lee B: Human immunodeficiency virus envelope (gp120) binding to DC-SIGN and primary dendritic cells is carbohydrate dependent but does not involve 2G12 or cyanovirin binding sites: implications for structural analyses of gp120-DC-SIGN binding. J Virol. 2002, 76 (24): 12855-12865. 10.1128/JVI.76.24.12855-12865.2002.PubMed CentralView ArticlePubMedGoogle Scholar
- Regnault A, Lankar D, Lacabanne V, Rodriguez A, Thery C, Rescigno M, Saito T, Verbeek S, Bonnerot C, Ricciardi-Castagnoli P, Amigorena S: Fcgamma receptor-mediated induction of dendritic cell maturation and major histocompatibility complex class I-restricted antigen presentation after immune complex internalization. J Exp Med. 1999, 189 (2): 371-380. 10.1084/jem.189.2.371.PubMed CentralView ArticlePubMedGoogle Scholar
- Morgan EL, Weigle WO: Polyclonal activation of murine B lymphocytes by Fc fragments. I. The requirement for two signals in the generation of the polyclonal antibody response induced by Fc fragments. J Immunol. 1980, 124 (3): 1330-1335.PubMedGoogle Scholar
- Obregon P, Chargelegue D, Drake PM, Prada A, Nuttall J, Frigerio L, Ma JK: HIV-1 p24-immunoglobulin fusion molecule: a new strategy for plant-based protein production. Plant Biotechnol J. 2006, 4 (2): 195-207. 10.1111/j.1467-7652.2005.00171.x.View ArticlePubMedGoogle Scholar
- Ma JK, Drake PM, Chargelegue D, Obregon P, Prada A: Antibody processing and engineering in plants, and new strategies for vaccine production. Vaccine. 2005, 23 (15): 1814-1818. 10.1016/j.vaccine.2004.11.011.View ArticlePubMedGoogle Scholar
- Canfield SM, Morrison SL: The binding affinity of human IgG for its high affinity Fc receptor is determined by multiple amino acids in the CH2 domain and is modulated by the hinge region. J Exp Med. 1991, 173 (6): 1483-1491. 10.1084/jem.173.6.1483.View ArticlePubMedGoogle Scholar
- Shields RL, Namenuk AK, Hong K, Meng YG, Rae J, Briggs J, Xie D, Lai J, Stadlen A, Li B, Fox JA, Presta LG: High resolution mapping of the binding site on human IgG1 for Fc gamma RI, Fc gamma RII, Fc gamma RIII, and FcRn and design of IgG1 variants with improved binding to the Fc gamma R. J Biol Chem. 2001, 276 (9): 6591-6604. 10.1074/jbc.M009483200.View ArticlePubMedGoogle Scholar
- Wang LX, Ni J, Singh S, Li H: Binding of high-mannose-type oligosaccharides and synthetic oligomannose clusters to human antibody 2G12: implications for HIV-1 vaccine design. Chem Biol. 2004, 11 (1): 127-134. 10.1016/S1074-5521(03)00299-0.PubMedGoogle Scholar
- Lee HK, Scanlan CN, Huang CY, Chang AY, Calarese DA, Dwek RA, Rudd PM, Burton DR, Wilson IA, Wong CH: Reactivity-based one-pot synthesis of oligomannoses: defining antigens recognized by 2G12, a broadly neutralizing anti-HIV-1 antibody. Angew Chem Int Ed Engl. 2004, 43 (8): 1000-1003. 10.1002/anie.200353105.View ArticlePubMedGoogle Scholar
- Law M, Cardoso RM, Wilson IA, Burton DR: Antigenic and immunogenic study of MPER-grafted gp120 antigens by DNA prime-protein boost immunization strategy. J Virol. 2007, 81 (8): 4272-4285. 10.1128/JVI.02536-06.PubMed CentralView ArticlePubMedGoogle Scholar
- Chapple SD, Jones IM: Non-polar distribution of green fluorescent protein on the surface of Autographa californica nucleopolyhedrovirus using a heterologous membrane anchor. J Biotechnol. 2002, 95 (3): 269-275. 10.1016/S0168-1656(02)00023-8.View ArticlePubMedGoogle Scholar
- Leonard CK, Spellman MW, Riddle L, Harris RJ, Thomas JN, Gregory TJ: Assignment of intrachain disulfide bonds and characterization of potential glycosylation sites of the type 1 recombinant human immunodeficiency virus envelope glycoprotein (gp120) expressed in Chinese hamster ovary cells. J Biol Chem. 1990, 265 (18): 10373-10382.PubMedGoogle Scholar
Copyright
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.