Multivalent interactions at biological interfaces occur frequently in nature and mediate recognition and interactions in essential physiological processes such as cell-to-cell adhesion. Multivalency is also a key principle that allows tight binding between pathogens and host cells during the initial stages of infection. One promising approach to prevent infection is the design of synthetic or semisynthetic multivalent binders that interfere with pathogen adhesion1,2,3,4. Here, we present a multivalent binder that is based on a spatially defined arrangement of ligands for the viral spike protein haemagglutinin of the influenza A virus. Complementary experimental and theoretical approaches demonstrate that bacteriophage capsids, which carry host cell haemagglutinin ligands in an arrangement matching the geometry of binding sites of the spike protein, can bind to viruses in a defined multivalent mode. These capsids cover the entire virus envelope, thus preventing its binding to the host cell as visualized by cryo-electron tomography. As a consequence, virus infection can be inhibited in vitro, ex vivo and in vivo. Such highly functionalized capsids present an alternative to strategies that target virus entry by spike-inhibiting antibodies5 and peptides6 or that address late steps of the viral replication cycle7.
Subscribe to Journal
Get full journal access for 1 year
only $15.58 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
The data supporting the findings of this study are available within the paper and its Supplementary Information. All relevant data are available from the authors upon reasonable request. Model coordinates of the HA–Sia binding pockets and the Qβ capsid surface are taken from the Protein Data Bank under accession numbers 1HGG and 1QBE, respectively.
Custom code is available from the corresponding author (S.L.).
Fasting, C. et al. Multivalency as a chemical organization and action principle. Angew. Chem. Int. Ed. 51, 10472–10498 (2012).
Gestwicki, J. E., Cairo, C. W., Strong, L. E., Oetjen, K. A. & Kiessling, L. L. Influencing receptor-ligand binding mechanisms with multivalent ligand architecture. J. Am. Chem. Soc. 124, 14922–14933 (2002).
Kiessling, L. L., Gestwicki, J. E. & Strong, L. E. Synthetic multivalent ligands in the exploration of cell-surface interactions. Curr. Opin. Chem. Biol. 4, 696–703 (2000).
Branson, T. R. et al. A protein-based pentavalent inhibitor of the cholera toxin B-subunit. Angew. Chem. Int. Ed. 53, 8323–8327 (2014).
Wu, N. C. & Wilson, I. A. Structural insights into the design of novel anti-influenza therapies. Nat. Struct. Mol. Biol. 25, 115–121 (2018).
Kadam, R. U. et al. Potent peptidic fusion inhibitors of influenza virus. Science 358, 496–502 (2017).
van de Wakker, S. I., Fischer, M. J. E. & Oosting, R. S. New drug-strategies to tackle viral-host interactions for the treatment of influenza virus infections. Eur. J. Pharm. 809, 178–190 (2017).
Weis, W. et al. Structure of the influenza virus haemagglutinin complexed with its receptor, sialic acid. Nature 333, 426–431 (1988).
Sauter, N. K. et al. Crystallographic detection of a second ligand binding site in influenza virus hemagglutinin. Proc. Natl Acad. Sci. USA 89, 324–328 (1992).
Stevens, J. et al. Structure of the uncleaved human H1 hemagglutinin from the extinct 1918 influenza virus. Science 303, 1866–1870 (2004).
Sauter, N. K. et al. Hemagglutinins from two influenza virus variants bind to sialic acid derivatives with millimolar dissociation constants: a 500-MHz proton nuclear magnetic resonance study. Biochemistry 28, 8388–8396 (1989).
Mammen, M., Choi, S. K. & Whitesides, G. M. Polyvalent Interactions in biological systems: implications for design and use of multivalent ligands and inhibitors. Angew. Chem. Int. Ed. 37, 2754–2794 (1998).
Lauster, D. et al. Multivalent peptide-nanoparticle conjugates for influenza-virus inhibition. Angew. Chem. Int. Ed. 56, 5931–5936 (2017).
Curk, T., Dobnikar, J. & Frenkel, D. Optimal multivalent targeting of membranes with many distinct receptors. Proc. Natl Acad. Sci. USA 114, 7210–7215 (2017).
Martinez-Veracoechea, F. J. & Frenkel, D. Designing super selectivity in multivalent nano-particle binding. Proc. Natl Acad. Sci. USA 108, 10963–10968 (2011).
Mammen, M., Dahmann, G. & Whitesides, G. M. Effective inhibitors of hemagglutination by influenza virus synthesized from polymers having active ester groups. Insight into mechanism of inhibition. J. Med. Chem. 38, 4179–4190 (1995).
Kwon, S. J. et al. Nanostructured glycan architecture is important in the inhibition of influenza A virus infection. Nat. Nanotechnol. 12, 48–54 (2017).
Kingery-Wood, J. E., Williams, K. W., Sigal, G. E. & Whitesides, G. M. The agglutination of erythrocytes by influenza virus is strongly inhibited by liposomes incorporating an analog of sialyl gangliosides. J. Am. Chem. Soc. 114, 7303–7305 (1992).
Wang, H. et al. Design and synthesis of glycoprotein-based multivalent glyco-ligands for influenza hemagglutinin and human galectin-3. Bioorg. Med. Chem. 21, 2037–2044 (2013).
Bandlow, V. et al. Spatial screening of hemagglutinin on influenza A virus particles: sialyl-LacNAc displays on DNA and PEG scaffolds reveal the requirements for bivalency enhanced interactions with weak monovalent binders. J. Am. Chem. Soc. 139, 16389–16397 (2017).
Waldmann, M. et al. A nanomolar multivalent ligand as entry inhibitor of the hemagglutinin of avian influenza. J. Am. Chem. Soc. 136, 783–788 (2014).
Strauch, E. M. et al. Computational design of trimeric influenza-neutralizing proteins targeting the hemagglutinin receptor binding site. Nat. Biotechnol. 35, 667–671 (2017).
Vonnemann, J. et al. Size dependence of steric shielding and multivalency effects for globular binding inhibitors. J. Am. Chem. Soc. 137, 2572–2579 (2015).
Liese, S. & Netz, R. R. Quantitative prediction of multivalent ligand-receptor binding affinities for influenza, cholera, and anthrax inhibition. ACS Nano 12, 4140–4147 (2018).
Yang, H. et al. Structure and receptor binding preferences of recombinant human A(H3N2) virus hemagglutinins. Virology 477, 18–31 (2015).
Ribeiro-Viana, R. et al. Virus-like glycodendrinanoparticles displaying quasi-equivalent nested polyvalency upon glycoprotein platforms potently block viral infection. Nat. Commun. 3, 1303 (2012).
Strable, E. et al. Unnatural amino acid incorporation into virus-like particles. Bioconjug. Chem. 19, 866–875 (2008).
Crich, D. & Li, W. α-Selective sialylations at –78 °C in nitrile solvents with a 1-adamantanyl thiosialoside. J. Org. Chem. 72, 7794–7797 (2007).
Hunter, C. A. & Anderson, H. L. What is cooperativity? Angew. Chem. Int. Ed. 48, 7488–7499 (2009).
Tate, M. D. et al. Playing hide and seek: how glycosylation of the influenza virus hemagglutinin can modulate the immune response to infection. Viruses 6, 1294–1316 (2014).
Roach, D. R. et al. Synergy between the host immune system and bacteriophage is essential for successful phage therapy against an acute respiratory pathogen. Cell Host Microbe 22, 38–47 (2017).
Budisa, N. et al. High-level biosynthetic substitution of methionine in proteins by its analogs 2-aminohexanoic acid, selenomethionine, telluromethionine and ethionine in Escherichia coli. Eur. J. Biochem 230, 788–796 (1995).
Desselberger, U. Relation of virus particle counts to the hemagglutinating activity of influenza virus suspensions measured by the HA pattern test and by use of the photometric HCU method. Arch. Virol. 49, 365–372 (1975).
Bhatia, S. et al. Linear polysialoside outperforms dendritic analogs for inhibition of influenza virus infection in vitro and in vivo. Biomaterials 138, 22–34 (2017).
Berg, J. et al. Tyk2 as a target for immune regulation in human viral/bacterial pneumonia. Eur. Respir. J. 50, 1601953 (2017).
Hocke, A. C. et al. Emerging human Middle East respiratory syndrome coronavirus causes widespread infection and alveolar damage in human lungs. Am. J. Respir. Crit. Care Med. 188, 882–886 (2013).
Weinheimer, V. K. Influenza A viruses target type II pneumocytes in the human lung. J. Infect. Dis. 206, 1685–1694 (2012).
Branston, S. D., Wright, J. & Keshavarz-Moore, E. A non-chromatographic method for the removal of endotoxins from bacteriophages. Biotechnol. Bioeng. 112, 1714–1719 (2015).
We thank Andrew K. Udit for providing the Qβ(K16M) plasmid and L. Artner for synthetic contributions. This work was supported by the German Research Foundation (DFG, SFB765, SPP1623 and SFB-TR84) and the Germany Ministry of Education and Research (BMBF, RAPID) as well as Charité 3R and the Einstein Foundation Berlin.
The authors declare no competing interests.
Peer review information Nature Nanotechnology thanks Andrew Ward and the other, anonymous, reviewers for their contribution to the peer review of this work.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Lauster, D., Klenk, S., Ludwig, K. et al. Phage capsid nanoparticles with defined ligand arrangement block influenza virus entry. Nat. Nanotechnol. 15, 373–379 (2020). https://doi.org/10.1038/s41565-020-0660-2