Sialic Acid-containing Receptors Research Articles

Expression of influenza A virus glycan receptor candidates in mallard, chicken, and tufted duck.

Influenza A virus (IAV) pandemics result from interspecies transmission events within the avian reservoir and further into mammals including humans. Receptor incompatibility due to differently expressed glycan structures between species has been suggested to limit zoonotic IAV transmission from the wild bird reservoir as well as between different bird species. Using glycoproteomics, we have studied the repertoires of expressed glycan structures with focus on putative sialic acid-containing glycan receptors for IAV in mallard, chicken and tufted duck; three bird species with different roles in the zoonotic ecology of IAV. The methodology used pinpoints specific glycan structures to specific glycosylation sites of identified glycoproteins and was also used to successfully discriminate α2-3- from α2-6-linked terminal sialic acids by careful analysis of oxonium ions released from glycopeptides in tandem MS/MS (MS2), and MS/MS/MS (MS3). Our analysis clearly demonstrated that all three bird species can produce complex N-glycans including α2-3-linked sialyl Lewis structures, as well as both N- and O- glycans terminated with both α2-3- and α2-6-linked Neu5Ac. We also found the recently identified putative IAV receptor structures, Man-6P N-glycopeptides, in all tissues of the three bird species. Furthermore, we found many similarities in the repertoires of expressed receptors both between the bird species investigated and to previously published data from pigs and humans. Our findings of sialylated glycan structures, previously anticipated to be mammalian specific, in all three bird species may have major implications for our understanding of the role of receptor incompatibility in interspecies transmission of IAV.

Examination of the Relationship between the ABO Blood Group and Susceptibility to SARS-CoV-2 Infection Risk

The ABO blood group system has been linked with multiple infectious and non-infectious diseases and disorders such as, hepatitis B, dengue haemorrhagic fever, cancer, cardiovascular diseases, hematologic disorders, metabolic diseases, malaria, etc. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is the virus that causes COVID-19 (Coronavirus disease 2019), a respiratory infectious disease that has become a global pandemic. Several reports have investigated the role of ABO blood groups in susceptibility/resistance to various infectious Diseases, and have proposed that ABO blood group polymorphism may be linked with COVID-19 susceptibility and clinical outcomes, however, the results were questionable. It has been suggested that blood type O may have a protective role against COVID-19 infection, as blood group O individuals were found COVID-19 positive in lower levels. This could emonstrate that those Individuals are less susceptible to infection, or are asymptomatic in higher proportions, or may be associated with a slightly lower risk for severe COVID-19 disease. It has been hypothesized that the mentioned association can be probably explained by the configuration of distribution of the sialic acid-containing receptors on host cell surfaces induced by ABO antigens through carbohydrate-carbohydrate interactions, that could maximize or minimize the SARS-CoV-2 virus spike protein (S) binding to the host cell. The classical viral entry through the ACE-2 receptors can be prevented by the anti-A antibodies that are produced from O and B blood group individuals. Experimental models based on cellular lines suggested a possible explanation for blood type configuration of infection showing that S protein/ACE-2-dependent adhesion to ACE 2-expressing cells was especially inhibited by natural or monoclonal human anti-A antibodies. Consequently, non-A blood groups individuals, mainly O, or B blood group, that produce anti-A antibodies, may have a lesser degree of susceptible to SARS-CoV-2 infection due to the inhibitory effects of anti-A antibodies. Therefore, it is possible that individuals with group A are more susceptible to SARS-CoV-2 infection and/or manifestation of a severe status.

Characterization of Influenza Virus Binding to Receptors on Isolated Cell Membranes.

An interplay between receptor-binding properties of influenza viruses (IVs) and spectrum of sialic acid-containing receptors on target cells in birds and mammals determine viral host range, tissue tropism, and pathogenicity. Here, we describe method that allows to characterize binding of IVs to biologically relevant cellular receptors using a conventional solid-phase enzyme-linked assay. In this method, we isolate plasma membranes from respiratory and intestinal epithelial cells of animal origin (Subheading 3.2). We adsorb the membranes in the wells of 96-well ELISA plates, incubate the membrane-coated wells with serially diluted IVs, and determine amounts of IVs attached to the membranes using viral ability to bind peroxidase-labeled sialoglycoprotein fetuin. Based on the concentration dependence of IV binding to the membrane, we estimate binding avidity and number of binding sites. We describe two variants of the assay in Subheadings 3.6 and 3.7 and provide examples.

Receptor-Binding Specificity of Influenza Viruses.

Influenza A virus infection begins with the attachment of virus particles to sialic acid-containing receptors on the surface of host cells. This attachment is mediated by the viral surface glycoprotein hemagglutinin (HA). Influenza A viruses have a wide host range, meaning they are able to infect many mammal and bird species. Influenza pandemics have been caused by viruses that contain genes from avian influenza viruses. Therefore, the infection of humans with avian influenza viruses, including avian H5Nx and H7Nx viruses, poses a huge threat to public health. These avian influenza viruses can transmit directly to humans from infected poultry, but do not spread easily among people, in part, due to differences in the receptor-binding specificities of human and avian influenza viruses. Therefore, conversion from avian- to human-type receptor-binding specificity is widely believed to be necessary for the efficient transmission of avian influenza viruses among humans. Accordingly, constant monitoring of the receptor-binding specificity of avian influenza viruses is important. In this chapter, we describe the protocol for assessing the receptor-binding specificity of influenza A viruses.

Hemagglutinin Stability and Its Impact on Influenza A Virus Infectivity, Pathogenicity, and Transmissibility in Avians, Mice, Swine, Seals, Ferrets, and Humans.

Genetically diverse influenza A viruses (IAVs) circulate in wild aquatic birds. From this reservoir, IAVs sporadically cause outbreaks, epidemics, and pandemics in wild and domestic avians, wild land and sea mammals, horses, canines, felines, swine, humans, and other species. One molecular trait shown to modulate IAV host range is the stability of the hemagglutinin (HA) surface glycoprotein. The HA protein is the major antigen and during virus entry, this trimeric envelope glycoprotein binds sialic acid-containing receptors before being triggered by endosomal low pH to undergo irreversible structural changes that cause membrane fusion. The HA proteins from different IAV isolates can vary in the pH at which HA protein structural changes are triggered, the protein causes membrane fusion, or outside the cell the virion becomes inactivated. HA activation pH values generally range from pH 4.8 to 6.2. Human-adapted HA proteins tend to have relatively stable HA proteins activated at pH 5.5 or below. Here, studies are reviewed that report HA stability values and investigate the biological impact of variations in HA stability on replication, pathogenicity, and transmissibility in experimental animal models. Overall, a stabilized HA protein appears to be necessary for human pandemic potential and should be considered when assessing human pandemic risk.

The influence of ABO blood groups on COVID-19 susceptibility and severity: A molecular hypothesis based on carbohydrate-carbohydrate interactions

The world is experiencing one of the most difficult moments in history with the COVID-19 pandemic, a disease caused by SARS-CoV-2, a new type of coronavirus. Virus infectivity is mediated by the binding of Spike transmembrane glycoprotein to specific protein receptors present on cell host surface. Spike is a homotrimer that emerges from the virion, each monomer containing two subunits named S1 and S2, which are related to cell recognition and membrane fusion, respectively. S1 is subdivided in domains S1A (or NTD) and S1B (or RBD), with experimental and in silico studies suggesting that the former binds to sialic acid-containing glycoproteins, such as CD147, whereas the latter binds to ACE2 receptor. Recent findings indicate that the ABO blood system modulates susceptibility and progression of infection, with type-A individuals being more susceptible to infection and/or manifestation of a severe condition. Seeking to understand the molecular mechanisms underlying this susceptibility, we carried out an extensive bibliographic survey on the subject. Based on this survey, we hypothesize that the correlation between the ABO blood system and susceptibility to SARS-CoV-2 infection can be presumably explained by the modulation of sialic acid-containing receptors distribution on host cell surface induced by ABO antigens through carbohydrate-carbohydrate interactions, which could maximize or minimize the virus Spike protein binding to the host cell. This model could explain previous sparse observations on the molecular mechanism of infection and can direct future research to better understand of COVID-19 pathophysiology.

The OAAH Family: Anti-Influenza Virus Lectins.

High mannose (HM)-binding Oscillatoria agardhii agglutinin homologue (OAAH) lectin family is an important class of anti-viral proteins. The OAAH family lectins show potent anti-influenza virus activity with EC50 of nanomolar levels by binding to HM glycans of the envelope glycoprotein hemagglutinin (HA), thereby inhibiting the viral entry into host cells. No broadly effective neutralizing vaccines for influenza virus are available due to the frequent antigenic drift caused by rapid mutations. Alternatives for vaccines need to be developed to prepare for a possible risk of future emergence of a highly virulent virus. Possible use of antiviral lectins is a simple and useful strategy to prevent viral infection by interfering with the interaction between viral HA and the host sialic acid-containing receptor. High-density glycans of surface HA are primary targets for the lectins to inhibit viral entry. In general, the anti-influenza virus potency of lectins is evaluated by a series of inhibitory assays for infection, such as neutral red dye uptake assay to determine the extent of viral cytopathic effect, and immunofluorescence microscopy to detect the expression of viral proteins in infected cells. Direct interaction between lectins and HA could be evaluated by enzyme-linked immunosorbent assay or surface plasmon resonance analysis.

Passage of influenza A/H3N2 viruses in human airway cells removes artefactual variants associated with neuraminidase-mediated binding.

Serological assays with modern influenza A/H3N2 viruses have become problematic due to the progressive reduction in the ability of viruses of this subtype to bind and agglutinate red blood cells (RBCs). This is due to reduced ability of the viral haemagglutinin (HA) glycoprotein to bind to the sialic acid-containing receptors presented by these cells. Additionally, as a result of reduced HA-mediated binding in cell culture, modern A/H3N2 viruses often acquire compensatory mutations during propagation that enable binding of cellular receptors through their neuraminidase (NA) surface protein. Viruses that have acquired this NA-mediated binding agglutinate RBCs through their NA, confusing the results of serological assays designed to assess HA antigenicity. Here we confirm with a large dataset that the acquisition of mutations that confer NA binding of RBCs is a culture artefact, and demonstrate that modern A/H3N2 isolates with acquired NA-binding mutations revert to a clinical-like NA sequence after a single passage in human airway epithelial (HAE) cells.

A Sialylated Voltage-Dependent Ca2+ Channel Binds Hemagglutinin and Mediates Influenza A Virus Entry into Mammalian Cells

Influenza A virus (IAV) infection is initiated by the attachment of the viral glycoprotein hemagglutinin (HA) to sialic acid on the host cell surface. However, the sialic acid-containing receptor crucial for IAV infection has remained unidentified. Here, we show that HA binds to the voltage-dependent Ca2+ channel Cav1.2 to trigger intracellular Ca2+ oscillations and subsequent IAV entry and replication. IAV entry was inhibited by Ca2+ channel blockers (CCBs) or by knockdown of Cav1.2. The CCB diltiazem also inhibited virus replication invivo. Reintroduction of wild-type but not the glycosylation-deficient mutants of Cav1.2 restored Ca2+ oscillations and virus infection in Cav1.2-depleted cells, demonstrating the significance of Cav1.2 sialylation. Taken together, we identify Cav1.2 as a sialylated host cell surface receptor that binds HA and is critical for IAV entry.

Glycan-binding preferences and genetic evolution of human seasonal influenza A(H3N2) viruses during 1999-2007 in Taiwan.

It is generally agreed that human influenza virus preferentially binds to α-2,6-linked sialic acid-containing receptors, and mutations that change the binding preference may alter virus infectivity and host tropism. Limited information is available on the glycan-binding specificity of epidemic influenza viruses. In this study, we systemically investigated the glycan-binding preferences of human influenza A(H3N2) viruses isolated from 1999 to 2007 in Taiwan using a high-throughput carbohydrate array. The binding patterns of 37 H3N2 viruses were classified into three groups with significant binding-pattern variations. The results showed that the carbohydrate-binding patterns of H3N2 varied over time. A phylogenetic analysis of the hemagglutinin gene also revealed progressive drift year to year. Of note, the viruses that caused large outbreaks in 1999 and 2003 showed glycan-binding preferences to both α-2,3 and α-2,6 sialylated glycans. Twenty amino acid substitutions were identified primarily at antigenic sites that might contribute to H3N2 virus evolution and the change in the glycan-binding patterns. This study provides not only a systematic analysis of the receptor-binding specificity of influenza clinical isolates but also information that could help to monitor the outbreak potential and virus evolution of influenza viruses.

Genetic Evidence for Erythrocyte Receptor Glycophorin B Expression Levels Defining a Dominant Plasmodium falciparum Invasion Pathway into Human Erythrocytes.

ABSTRACTPlasmodium falciparum, the parasite that causes the deadliest form of malaria, has evolved multiple proteins known as invasion ligands that bind to specific erythrocyte receptors to facilitate invasion of human erythrocytes. The EBA-175/glycophorin A (GPA) and Rh5/basigin ligand-receptor interactions, referred to as invasion pathways, have been the subject of intense study. In this study, we focused on the less-characterized sialic acid-containing receptors glycophorin B (GPB) and glycophorin C (GPC). Through bioinformatic analysis, we identified extensive variation in glycophorin B (GYPB) transcript levels in individuals from Benin, suggesting selection from malaria pressure. To elucidate the importance of the GPB and GPC receptors relative to the well-described EBA-175/GPA invasion pathway, we used an ex vivo erythrocyte culture system to decrease expression of GPA, GPB, or GPC via lentiviral short hairpin RNA transduction of erythroid progenitor cells, with global surface proteomic profiling. We assessed the efficiency of parasite invasion into knockdown cells using a panel of wild-type P. falciparum laboratory strains and invasion ligand knockout lines, as well as P. falciparum Senegalese clinical isolates and a short-term-culture-adapted strain. For this, we optimized an invasion assay suitable for use with small numbers of erythrocytes. We found that all laboratory strains and the majority of field strains tested were dependent on GPB expression level for invasion. The collective data suggest that the GPA and GPB receptors are of greater importance than the GPC receptor, supporting a hierarchy of erythrocyte receptor usage in P. falciparum.

Neuraminidase activity of blue eye disease porcine rubulavirus: Specificity, affinity and inhibition studies

Porcine rubulavirus (PorPV), also known as La Piedad Michoacan Virus (LPMV) causes encephalitis and reproductive failure in newborn and adult pigs, respectively. The hemagglutinin–neuraminidase (HN) glycoprotein is the most exposed and antigenic of the virus proteins. HN plays central roles in PorPV infection; i.e., it recognizes sialic acid-containing cell receptors that mediate virus attachment and penetration; in addition, its neuraminidase (sialic acid releasing) activity has been proposed as a virulence factor.This work describes the purification and characterization of PorPV HN protein (isolate PAC1). The specificity of neuraminidase is restricted to sialyl(α2,3)lactose (3SL). HN showed typical Michaelis-Menten kinetics with fetuin as substrate (km=0.029μM, Vmax=522.8nmolmin−1mg−1). When 3SL was used as substrate, typical cooperative kinetics were found (S50=0.15μM, Vmax=154.3nmolmin−1mg−1). The influenza inhibitor zanamivir inhibited the PorPV neuraminidase with IC50 of 0.24μM. PorPV neuraminidase was activated by Ca2+ and inhibited by nucleoside triphosphates with the level of inhibition depending on phosphorylation level. The present results open possibilities to study the role of neuraminidase in the pathogenicity of PorPV infection and its potential inhibitors.

Recent H3N2 Viruses Have Evolved Specificity for Extended, Branched Human-type Receptors, Conferring Potential for Increased Avidity.

Human and avian influenza viruses recognize different sialic acid-containing receptors, referred to as human-type (NeuAcα2-6Gal) and avian-type (NeuAcα2-3Gal), respectively. This presents a species barrier for aerosol droplet transmission of avian viruses in humans and ferrets. Recent reports have suggested that current human H3N2 viruses no longer have strict specificity toward human-type receptors. Using an influenza receptor glycan microarray with extended airway glycans, we find that H3N2 viruses have in fact maintained human-type specificity, but they have evolved preference for a subset of receptors comprising branched glycans with extended poly-N-acetyl-lactosamine (poly-LacNAc) chains, a specificity shared with the 2009 pandemic H1N1 (Cal/04) hemagglutinin. Lipid-linked versions of extended sialoside receptors can restore susceptibility of sialidase-treated MDCK cells to infection byboth recent (A/Victoria/361/11) and historical (A/Hong Kong/8/1968) H3N2 viruses. Remarkably, these human-type receptors with elongated branches have the potential to increase avidity by simultaneously binding to two subunits of a single hemagglutinin trimer.

A DNA Aptamer Against Influenza A Virus: An Effective Inhibitor to the Hemagglutinin-Glycan Interactions.

Most therapeutical nucleic acid aptamers tend to inhibit protein-protein interactions and thereby function as antagonists. Attachment of the influenza virus surface glycoprotein hemagglutinin (HA) to sialic acid-containing host cell receptors (glycan) facilitates the initial stage of viral infection. Inhibition of the attachment may result in an antiviral effect on the proliferation of the influenza virus. To develop therapeutically interesting agents, we selected two single-stranded DNA (ssDNA) aptamers specific to the HA protein of H1N1 influenza virus (A/Puerto Rico/8/1934) through a procedure of systematic evolution of ligands by exponential enrichment. As it showed a higher binding affinity for HA protein (Kd = 78 ± 1 nM), aptamer 1 was tested for its ability to interfere with HA-glycan interactions using chicken red blood cell hemagglutination and microneutralization assays, which demonstrated that it significantly suppressed the viral infection in host cells. These results indicate that the isolated ssDNA aptamer may be developed as an antiviral agent against influenza through appropriate therapeutic formulation.

What adaptive changes in hemagglutinin and neuraminidase are necessary for emergence of pandemic influenza virus from its avian precursor?

Wild ducks serve as the primary host for numerous and various influenza type A viruses. Occasionally, viruses from this reservoir can be transferred to other host species and cause outbreaks of influenza in fowl, swine, and horses, as well as result in novel human pandemics. Cellular tropism and range of susceptible host species are determined by interaction between virus and receptor molecules on cells. Here we discuss modern data regarding molecular features underlying interactions of influenza viruses with cellular receptors as well as a role for receptor specificity in interspecies transmission. By analyzing the earliest available pandemic influenza viruses (1918, 1957, 1968, 2009), we found that hemagglutinin reconfigured to recognize 2-6 sialic acid-containing receptors in the human upper airway tract together with altered enzymatic activity of neuraminidase necessary for maintaining functional balance with hemagglutinin are responsible for effective spread of influenza viruses in human populations. Resistance to low pH also contributes to this. Thus, a combination of such parameters makes it possible that influenza viruses give rise to novel pandemics.

Microsecond-Resolution Recording of T4 Lysozyme Observes a Brownian Ratchet

Microsecond-Resolution Recording of T4 Lysozyme Observes a Brownian Ratchet

Nothing to Sneeze at: A Full-Scale Computational Model of the Human Influenza Virion

Tackling the ongoing challenge of influenza infectivity would benefit greatly from a molecular understanding of why the influenza A virion exhibits seasonal patterns of infectivity and has wide-ranging survival times in different environments. A computational approach to the study of the behaviour of the virion that focuses on the poorly-understood structural and dynamic role of the lipids is presented here. We have combined experimental data from X-ray crystallography, NMR spectroscopy, cryoelectron microscopy, and lipidomics to build a full-scale computational model of the influenza A virion. This represents the first set of microsecond-scale molecular dynamics simulations of an enveloped virion in explicit solvent that we are aware of. We report results for a set of simulations at different temperatures and with varying lipid compositions. The Forssman glycolipid, which is prevalent in the influenza A lipidome, influences several biophysical characteristics of the virion model, including diffusion and clustering of proteins and lipids. The distribution of peplomers on the virion surface is consistent with accessibility to bivalent antibodies. The distances of binding sites for host cell sialic acid-containing receptors have been analyzed in the virion model for a variety of planar host cell membrane attack orientations. The relatively rigid membrane of the influenza A virion model exhibits a number of biophysical properties consistent with experimental measurements, and may serve as a useful platform for in silico assessment of virion behaviour.

Sialic acid-binding protein Sp2CBMTD protects mice against lethal challenge with emerging influenza A (H7N9) virus.

Compounds that target the cellular factors essential for influenza virus replication represent an innovative approach to antiviral therapy. Sp2CBMTD is a genetically engineered multivalent protein that masks sialic acid-containing cellular receptors on the respiratory epithelium, which are recognized by influenza viruses. Here, we evaluated the antiviral potential of Sp2CBMTD against lethal infection in mice with an emerging A/Anhui/1/2013 (H7N9) influenza virus and addressed the mechanistic basis of its activity in vivo. Sp2CBMTD was administered to mice intranasally as a single or repeated dose (0.1, 1, 10, or 100 μg) before (day -7, -3, and/or -1) or after (6 or 24 h) H7N9 virus inoculation. A single Sp2CBMTD dose (10 or 100 μg) protected 80% to 100% of the mice when administered 7 days before the H7N9 lethal challenge. Repeated Sp2CBMTD administration conferred the highest protection, resulting in 100% survival of the mice even at the lowest dose tested (0.1 μg). When treatment began 24 h after exposure to the H7N9 virus, a single administration of 100 μg of Sp2CBMTD protected 40% of the mice from death. The administration of Sp2CBMTD induced the pulmonary expression of proinflammatory mediators (interleukin-6 [IL-6], IL-1β, RANTES, monocyte chemotactic protein-1 [MCP-1], macrophage inflammatory protein-1α [MIP-1α], and inducible protein [IP-10]) and recruited neutrophils to the respiratory tract before H7N9 virus infection, which resulted in less pronounced inflammation and rapid virus clearance from mouse lungs. Sp2CBMTD administration did not affect the virus-specific adaptive immune response, which was sufficient to protect against reinfection with a higher dose of homologous H7N9 virus or heterologous H5N1 virus. Thus, Sp2CBMTD was effective in preventing H7N9 infections in a lethal mouse model and holds promise as a prophylaxis option against zoonotic influenza viruses.

Influenza virus hemagglutinin as a vaccine antigen produced in bacteria.

Recombinant subunit vaccines based on hemagglutinin proteins produced in bacteria (bacterial HAs) are promising candidates for enhancing the supply of vaccines against influenza, especially for a pandemic. Over 20 years after the failure to obtain the antigen with native HA characteristics in the early 1980's, there are increasing data on successful production of HA proteins in bacteria. The vast majority of bacterial HAs have been based on the HA1 subunit of HA expressed separately or as a component of conjugate vaccines, but those based on the ectodomain and the HA2 subunit have also been reported. The most of HAs have been efficiently expressed as insoluble aggregates called inclusion bodies. Refolded and purified proteins were extensively studied for structure, the ability to bind to sialic acid-containing receptors, antigenicity, immunogenicity and efficacy. The results from these studies contradict the view that glycosylation determines the correct structure of the hemagglutinin, as they proved that bacterial HAs can be valuable vaccine antigens when appropriate folding and purification methods are applied to rationally designed proteins. The best evidence for success in bacterial production of protective HA is that vaccines based on proprietary Toll-like Receptor (VaxInnate) and bacteriophage Qβ-VLPs (Cytos Biotechnology) technologies have been advanced to clinical studies.

Structure analysis of the major capsid proteins of human polyomaviruses 6 and 7 reveals an obstructed sialic acid binding site.

Human polyomavirus 6 (HPyV6) and HPyV7 are commonly found on human skin. We have determined the X-ray structures of their major capsid protein, VP1, at resolutions of 1.8 and 1.7 Å, respectively. In polyomaviruses, VP1 commonly determines antigenicity as well as cell-surface receptor specificity, and the protein is therefore linked to attachment, tropism, and ultimately, viral pathogenicity. The structures of HPyV6 and HPyV7 VP1 reveal uniquely elongated loops that cover the bulk of the outer virion surfaces, obstructing a groove that binds sialylated glycan receptors in many other polyomaviruses. In support of this structural observation, interactions of VP1 with α2,3- and α2,6-linked sialic acids could not be detected in solution by nuclear magnetic resonance spectroscopy. Single-cell binding studies indicate that sialylated glycans are likely not required for initial attachment to cultured human cells. Our findings establish distinct antigenic properties of HPyV6 and HPyV7 capsids and indicate that these two viruses engage nonsialylated receptors. Eleven new human polyomaviruses, including the skin viruses HPyV6 and HPyV7, have been identified during the last decade. In contrast to better-studied polyomaviruses, the routes of infection, cell tropism, and entry pathways of many of these new viruses remain largely mysterious. Our high-resolution X-ray structures of major capsid proteins VP1 from HPyV6 and from HPyV7 reveal critical differences in surface morphology from those of all other known polyomavirus structures. A groove that engages specific sialic acid-containing glycan receptors in related polyomaviruses is obstructed, and VP1 of HPyV6 and HPyV7 does not interact with sialylated compounds in solution or on cultured human cells. A comprehensive comparison with other structurally characterized polyomavirus VP1 proteins enhances our understanding of molecular determinants that underlie receptor specificity, antigenicity, and, ultimately, pathogenicity within the polyomavirus family and highlight the need for structure-based analysis to better define phylogenetic relationships within the growing polyomavirus family and perhaps also for other viruses.