Structural Comparison of Different Antibodies Interacting with Parvovirus Capsids

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Parvovirus capsids are small but complex molecular machines responsible for undertaking many of the steps of cell infection, genome packing, and cell-tocell as well as host-to-host transfer. The details of parvovirus infection of cells are still not fully understood, but the processes must involve small changes in the capsid structure that allow the endocytosed virus to escape from the endosome, pass through the cell cytoplasm, and deliver the single-stranded DNA (ssDNA) genome to the nucleus, where viral replication occurs. Here, we examine capsid substitutions that eliminate canine parvovirus (CPV) infectivity and identify how those mutations changed the capsid structure or altered interactions with the infectious pathway. Amino acid substitutions on the exterior surface of the capsid (Gly299Lys/Ala300Lys) altered the binding of the capsid to transferrin receptor type 1 (TfR), particularly during virus dissociation from the receptor, but still allowed efficient entry into both feline and canine cells without successful infection. These substitutions likely control specific capsid structural changes resulting from TfR binding required for infection. A second set of changes on the interior surface of the capsid reduced viral infectivity by Ͼ100-fold and included two cysteine residues and neighboring residues. One of these substitutions, Cys270Ser, modulates a VP2 cleavage event found in ϳ10% of the capsid proteins that also was shown to alter capsid stability. A neighboring substitution, Pro272Lys, significantly reduced capsid assembly, while a Cys273Ser change appeared to alter capsid transport from the nucleus. These mutants reveal additional structural details that explain cell infection processes of parvovirus capsids.IMPORTANCE Parvoviruses are commonly found in both vertebrate and invertebrate animals and cause widespread disease. They are also being developed as oncolytic therapeutics and as gene therapy vectors. Most functions involved in infection or transduction are mediated by the viral capsid, but the structure-function correlates of the capsids and their constituent proteins are still incompletely understood, especially in relation to identifying capsid processes responsible for infection and release from the cell. Here, we characterize the functional effects of capsid protein mutations that result in the loss of virus infectivity, giving a better understanding of the portions of the capsid that mediate essential steps in successful infection pathways and how they contribute to viral infectivity.

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Parvovirus Capsid Structures Required for Infection: Mutations Controlling Receptor Recognition and Protease Cleavages

Callaway

Feng

Lee

et al. 2017

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“…The X-ray crystal structures of EV71 strains MY104 (5) and Fuyang (6) were fitted separately into the asymmetric virus-Fab cryo-EM reconstruction by superimposing symmetry elements. The crystal structure of a murine antibody Fab (PDB accession code 3GK8) (29) was fitted into the Fab density to identify contacts on the virus surface (Fig. 3B) (PDB accession code 3J3Z).…”

Section: Resultsmentioning

confidence: 99%

“…Fitting of the atomic structure of murine antibody Fab 3GK8 (29) was done first by eye and then refined in UCSF Chimera (28). The refined fit of the Fab (correlation coefficient value of 0.743) was used to identify all atoms within 4 Å of the fitted virus structures to generate the Fab-binding footprint.…”

Section: Methodsmentioning

confidence: 99%

A Strain-Specific Epitope of Enterovirus 71 Identified by Cryo-Electron Microscopy of the Complex with Fab from Neutralizing Antibody

Lee

Cifuente

Ashley

et al. 2013

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e Enterovirus 71 (EV71) is a picornavirus that causes outbreaks of hand, foot, and mouth disease (HFMD), primarily in the Asia-Pacific area. Unlike coxsackievirus A16, which also causes HFMD, EV71 induces severe neuropathology leading to high fatalities, especially among children under the age of 6 years. Currently, no established vaccines or treatments are available against EV71 infection. The monoclonal antibody MA28-7 neutralizes only specific strains of EV71 that have a conserved glycine at amino acid VP1-145, a surface-exposed residue that maps to the 5-fold vertex and that has been implicated in receptor binding. The cryo-electron microscopy structure of a complex between EV71 and the Fab fragment of MA28-7 shows that only one Fab fragment occupies each 5-fold vertex. A positively charged patch, which has also been implicated in receptor binding, lies within the Fab footprint. We identify the strain-specific epitope of EV71 and discuss the possible neutralization mechanisms of the antibody. E nterovirus 71 (EV71) infection causes outbreaks of hand, foot, and mouth disease (HFMD), predominantly in the Asia-Pacific region (1, 2). Whereas most EV71 infections are self-limiting, neurological and systemic complications can develop that range from aseptic meningitis to encephalitis and acute flaccid paralysis. Infection can lead to lethal pulmonary edema and heart failure (2), with mortality being especially high in young children under the age of 6 (2, 3). As seasonal outbreaks of HFMD are recurring around the world, development of a vaccine and antiviral therapies for EV71 has become an urgent concern.A member of the Picornaviridae family, EV71 has a nonenveloped, icosahedral capsid comprised of 60 copies of each of four viral structural proteins (VP1 to VP4) (4). Recent studies have solved the structures for three strains of EV71 (MY104 [5], Fuyang [6], and 1095 [7]), demonstrating that EV71 has the general features of picornavirus capsids, including the 5-fold "mesa" and the depression around the mesa called the "canyon" (5-8). Conserved residues VP1-242K and VP1-244K form positively charged patches on the 5-fold mesa (6), and this symmetry-related clustering of positive charges has been suggested as a common mechanism for heparan sulfate binding in enteroviruses (9).Several cellular receptors for EV71 have been identified: scavenger receptor B2 (SCARB2), P-selectin glycoprotein ligand-1 (PSGL-1), and heparan sulfate (HS) (10-12). SCARB2, which is expressed on a broad variety of cell types, likely binds to the virus canyon and induces the transition of the virion that is required for uncoating (13-15). PSGL-1, which is expressed exclusively on lymphocytes, binds only specific EV71 strains and supports viral replication in lymphocytes in a PSGL-1-dependent manner (11). According to recent studies, PSGL-1 and HS bind the positively charged patches on the 5-fold mesa of EV71 and provide initial attachment on the cell (12,16,17). We recently found that the PSGL-1 binding phenotype of EV71 strains is regulated b...

“…These four mutations map to or near the capsid surface and influence infection by altering binding to the carnivore transferrin receptor (TfR), the host cell attachment protein for these viruses (5). Additionally, these four mutations have been shown to alter antibody binding, as they cluster to a position on the capsid surface where an antigenic site overlaps with the receptor-binding site (5)(6)(7)(8). Besides these four mutations, there are other changes seen between CPV-2 and later isolates that became globally distributed.…”

mentioning

confidence: 99%

Global Displacement of Canine Parvovirus by a Host-Adapted Variant: Structural Comparison between Pandemic Viruses with Distinct Host Ranges

Organtini

Allison

Lukk

et al. 2015

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c Canine parvovirus type 2 (CPV-2) emerged in 1978 and spread worldwide within 2 years. Subsequently, CPV-2 was completely replaced by the variant CPV-2a, which is characterized by four specific capsid (VP2) mutations. The X-ray crystal structure of the CPV-2a capsid shows that each mutation confers small local changes. The loss of a hydrogen bond and introduction of a glycine residue likely introduce flexibility to sites that control interactions with the host receptor, antibodies, and sialic acids.