Towards conformational fidelity of a quaternary HIV-1 epitope: computational design and directed evolution of a minimal V1V2 antigen

Abstract Structure-based approaches to antigen design utilize insights from antibody (Ab):antigen interactions and a refined understanding of protective Ab responses to engineer novel antigens presenting epitopes with conformations relevant to eliciting or discovering protective humoral responses. For human immunodeficiency virus-1 (HIV-1), one model of protection is provided by broadly neutralizing Abs (bnAbs) against epitopes present in the closed prefusion trimeric conformation of HIV-1 envelope glycoprotein, such as the variable loops 1–2 (V1V2) apex. Here, computational design and directed evolution yielded a novel V1V2 sequence variant with potential utility for inclusion in an immunogen for eliciting bnAbs, or as an epitope probe for their detection. The computational design goal was to engineer a minimal single-chain antigen with three copies of the V1V2 loops to support maintenance of closed prefusion V1V2 trimeric conformation and presentation of bnAb epitopes. Via directed evolution of this computationally designed single-chain antigen, we isolated a V1V2 sequence variant that in monomeric form exhibited preferential recognition by quaternary-preferring and conformation-dependent mAbs. Structural context and transferability of this phenotype to V1V2 sequences from all strains of HIV-1 tested suggest a conformation-stabilizing effect. This example demonstrates the potential utility of computational design and directed evolution-based protein engineering strategies to develop minimal, conformation-stabilized epitope-specific antigens.

[1]  K. Wittrup,et al.  Highly avid magnetic bead capture: An efficient selection method for de novo protein engineering utilizing yeast surface display , 2009, Biotechnology progress.

[2]  David Baker,et al.  Proof of principle for epitope-focused vaccine design , 2014, Nature.

[3]  Alexandre G. de Brevern,et al.  Protein Peeling 3D: new tools for analyzing protein structures , 2011, Bioinform..

[4]  D. Baker,et al.  RosettaRemodel: A Generalized Framework for Flexible Backbone Protein Design , 2011, PloS one.

[5]  Adrian Apetri,et al.  A stable trimeric influenza hemagglutinin stem as a broadly protective immunogen , 2015, Science.

[6]  S. Zolla-Pazner,et al.  Rationally Designed Immunogens Targeting HIV-1 gp120 V1V2 Induce Distinct Conformation-Specific Antibody Responses in Rabbits , 2016, Journal of Virology.

[7]  Allan C. deCamp,et al.  HIV-1 Envelope Glycoproteins from Diverse Clades Differentiate Antibody Responses and Durability among Vaccinees , 2018, Journal of Virology.

[8]  Karen G. Dowell,et al.  Multiplexed Fc array for evaluation of antigen-specific antibody effector profiles , 2017, Journal of immunological methods.

[9]  Jerome H. Kim,et al.  Vaccination with ALVAC and AIDSVAX to prevent HIV-1 infection in Thailand. , 2009, The New England journal of medicine.

[10]  Guido Ferrari,et al.  Immune-correlates analysis of an HIV-1 vaccine efficacy trial. , 2012, The New England journal of medicine.

[11]  John P. Moore,et al.  Crystal Structure of a Soluble Cleaved HIV-1 Envelope Trimer , 2013, Science.

[12]  A. Pinter,et al.  Presentation of native epitopes in the V1/V2 and V3 regions of human immunodeficiency virus type 1 gp120 by fusion glycoproteins containing isolated gp120 domains , 1994, Journal of virology.

[13]  E. Go,et al.  Glycosylation site-specific analysis of HIV envelope proteins (JR-FL and CON-S) reveals major differences in glycosylation site occupancy, glycoform profiles, and antigenic epitopes' accessibility. , 2008, Journal of proteome research.

[14]  K Dane Wittrup,et al.  Isolating and engineering human antibodies using yeast surface display , 2006, Nature Protocols.

[15]  Gevorg Grigoryan,et al.  Rapid search for tertiary fragments reveals protein sequence–structure relationships , 2015, Protein science : a publication of the Protein Society.

[16]  Baoshan Zhang,et al.  Structural basis for diverse N-glycan recognition by HIV-1–neutralizing V1–V2–directed antibody PG16 , 2013, Nature Structural &Molecular Biology.

[17]  D. Baker,et al.  Computation-Guided Backbone Grafting of a Discontinuous Motif onto a Protein Scaffold , 2011, Science.

[18]  John P. Moore,et al.  Asymmetric recognition of the HIV-1 trimer by broadly neutralizing antibody PG9 , 2013, Proceedings of the National Academy of Sciences.

[19]  Raphael Gottardo,et al.  Plasma IgG to Linear Epitopes in the V2 and V3 Regions of HIV-1 gp120 Correlate with a Reduced Risk of Infection in the RV144 Vaccine Efficacy Trial , 2013, PloS one.

[20]  J. Sodroski,et al.  Human anti-V2 monoclonal antibody that neutralizes primary but not laboratory isolates of human immunodeficiency virus type 1 , 1994, Journal of virology.

[21]  Pham Phung,et al.  Broad and Potent Neutralizing Antibodies from an African Donor Reveal a New HIV-1 Vaccine Target , 2009, Science.

[22]  Pham Phung,et al.  Broad neutralization coverage of HIV by multiple highly potent antibodies , 2011, Nature.

[23]  C. Bailey-Kellogg,et al.  High-throughput, multiplexed IgG subclassing of antigen-specific antibodies from clinical samples. , 2012, Journal of immunological methods.

[24]  Serge A. Hazout,et al.  'Protein Peeling': an approach for splitting a 3D protein structure into compact fragments , 2006, Bioinform..

[25]  John P. Moore,et al.  Stabilization of the Soluble, Cleaved, Trimeric Form of the Envelope Glycoprotein Complex of Human Immunodeficiency Virus Type 1 , 2002, Journal of Virology.

[26]  Young Do Kwon,et al.  Structure of HIV-1 gp120 V1/V2 domain with broadly neutralizing antibody PG9 , 2011, Nature.

[27]  Fragments of the V1/V2 domain of HIV-1 glycoprotein 120 engineered for improved binding to the broadly neutralizing PG9 antibody. , 2016, Molecular immunology.

[28]  Eric T. Boder,et al.  Yeast surface display for screening combinatorial polypeptide libraries , 1997, Nature Biotechnology.

[29]  R. Wyatt,et al.  Cleavage-independent HIV-1 Env trimers engineered as soluble native spike mimetics for vaccine design. , 2015, Cell reports.

[30]  S. Zolla-Pazner,et al.  Functional and immunochemical cross-reactivity of V2-specific monoclonal antibodies from HIV-1-infected individuals. , 2012, Virology.

[31]  D. van der Spoel,et al.  GROMACS: A message-passing parallel molecular dynamics implementation , 1995 .

[32]  Timothy Cardozo,et al.  Structure–function relationships of HIV-1 envelope sequence-variable regions refocus vaccine design , 2010, Nature Reviews Immunology.

[33]  J. Mascola,et al.  Serotyping of primary human immunodeficiency virus type 1 isolates from diverse geographic locations by flow cytometry , 1995, Journal of virology.

[34]  William R Schief,et al.  Advances in structure-based vaccine design. , 2013, Current opinion in virology.

[35]  M. Ackerman,et al.  Directed Evolution of a Yeast-Displayed HIV-1 SOSIP gp140 Spike Protein toward Improved Expression and Affinity for Conformational Antibodies , 2015, PloS one.

[36]  R. Rappuoli,et al.  Reverse vaccinology 2.0: Human immunology instructs vaccine antigen design , 2016, The Journal of experimental medicine.

[37]  Gary J. Nabel,et al.  Vaccine-Induced IgG Antibodies to V1V2 Regions of Multiple HIV-1 Subtypes Correlate with Decreased Risk of HIV-1 Infection , 2014, PloS one.

[38]  Cinque S. Soto,et al.  Structure-Based Design of a Fusion Glycoprotein Vaccine for Respiratory Syncytial Virus , 2013, Science.

[39]  What mAbs tell us about shapes: multiple roads lead to Rome. , 2013, Immunity.

[40]  Jonathan R. McDaniel,et al.  Structures of HIV-1-Env V1V2 with broadly neutralizing antibodies reveal commonalities that enable vaccine design , 2015, Nature Structural &Molecular Biology.

[41]  Mario Roederer,et al.  Rational Design of Envelope Identifies Broadly Neutralizing Human Monoclonal Antibodies to HIV-1 , 2010, Science.

[42]  John P. Moore,et al.  A Next-Generation Cleaved, Soluble HIV-1 Env Trimer, BG505 SOSIP.664 gp140, Expresses Multiple Epitopes for Broadly Neutralizing but Not Non-Neutralizing Antibodies , 2013, PLoS pathogens.

[43]  S. Zolla-Pazner,et al.  Prevalence of a V2 epitope in clade B primary isolates and its recognition by sera from HIV-1-infected individuals. , 1997, AIDS.

[44]  John P. Moore,et al.  Recombinant HIV envelope trimer selects for quaternary-dependent antibodies targeting the trimer apex , 2014, Proceedings of the National Academy of Sciences.

[45]  P. Plateau,et al.  Direct random mutagenesis of gene-sized DNA fragments using polymerase chain reaction. , 1995, Analytical biochemistry.

[46]  D. Burton,et al.  Presenting native-like trimeric HIV-1 antigens with self-assembling nanoparticles , 2016, Nature Communications.

[47]  J. Sodroski,et al.  Solid-Phase Proteoliposomes Containing Human Immunodeficiency Virus Envelope Glycoproteins , 2002, Journal of Virology.

[48]  Jerome H. Kim,et al.  Antibody-Dependent Cellular Cytotoxicity-Mediating Antibodies from an HIV-1 Vaccine Efficacy Trial Target Multiple Epitopes and Preferentially Use the VH1 Gene Family , 2012, Journal of Virology.

[49]  Guido Ferrari,et al.  Vaccine induction of antibodies against a structurally heterogeneous site of immune pressure within HIV-1 envelope protein variable regions 1 and 2. , 2013, Immunity.