Glycosylation at hemagglutinin Asn-167 protects the H6N1 avian influenza virus from tryptic cleavage at Arg-201 and maintains the viral infectivity

He, Jijun; Chiu, Yi-Chung; Chang, Shih‐Chung; Wang, Ching-Ho; Juang, Rong-Huay

doi:10.1016/j.virusres.2014.12.010

Cited by 7 publications

(4 citation statements)

References 35 publications

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“…In the structures, we observed five glycans (at HA 1 21, 33, 169, 291 and HA 2 154) for both H6 HAs ( Fig 1A and 1B ). The glycan at HA 1 169 is located at the HA 1 -HA 1 monomer interface, which was reported to protect H6 HA from trypsin cleavage at R201 [ 42 ]. The overall structures of GD and TW H6 HAs are very similar, where the root-mean-square deviation (RMSD) for all Cα atoms is at 0.67 Å ( Fig 1C ).…”

Section: Resultsmentioning

confidence: 99%

Structural and Functional Studies of Influenza Virus A/H6 Hemagglutinin

Kondrashkina

Wang

2015

PLoS ONE

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In June 2013, the first human infection by avian influenza A(H6N1) virus was reported in Taiwan. This incident raised the concern for possible human epidemics and pandemics from H6 viruses. In this study, we performed structural and functional investigation on the hemagglutinin (HA) proteins of the human-infecting A/Taiwan/2/2013(H6N1) (TW H6) virus and an avian A/chicken/Guangdong/S1311/2010(H6N6) (GD H6) virus that transmitted efficiently in guinea pigs. Our results revealed that in the presence of HA1 Q226, the triad of HA1 S137, E190 and G228 in GD H6 HA allows the binding to both avian- and human-like receptors with a slight preference for avian receptors. Its conservation among the majority of H6 HAs provides an explanation for the broader host range of this subtype. Furthermore, the triad of N137, V190 and S228 in TW H6 HA may alleviate the requirement for a hydrophobic residue at HA1 226 of H2 and H3 HAs when binding to human-like receptors. Consequently, TW H6 HA has a slight preference for human receptors, thus may represent an intermediate towards a complete human adaptation. Importantly, the triad observed in TW H6 HA is detected in 74% H6 viruses isolated from Taiwan in the past 14 years, suggesting an elevated threat of H6 viruses from this region to human health. The novel roles of the triad at HA1 137, 190 and 228 of H6 HA in binding to receptors revealed here may also be used by other HA subtypes to achieve human adaptation, which needs to be further tested in laboratory and closely monitored in field surveillance.

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Section: Resultsmentioning

confidence: 99%

Structural and Functional Studies of Influenza Virus A/H6 Hemagglutinin

Kondrashkina

Wang

2015

PLoS ONE

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“…In particular, IAV strains undergo amino acid mutations to accumulate new N -glycosylation sequons during seasonal circulation in the human population (1–4). Thus, based primarily on genetic sequencing evidence (5–7), glycans appear to shield HA antigenic sites from host antibody recognition, as part of the evolutionary antigenic drift (8–11). Although this putative shielding effect of glycans apparently improves the ability of the virus to escape antibody neutralization, the virus must compensate for loss in receptor avidity from steric interference caused by the glycans proximal to the sialic acid binding sites on the HA head (12, 13).…”

mentioning

confidence: 99%

Integrated Omics and Computational Glycobiology Reveal Structural Basis for Influenza A Virus Glycan Microheterogeneity and Host Interactions

Khatri

Klein

White

et al. 2016

Molecular & Cellular Proteomics

114

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Despite sustained biomedical research effort, influenza A virus remains an imminent threat to the world population and a major healthcare burden. The challenge in developing vaccines against influenza is the ability of the virus to mutate rapidly in response to selective immune pressure. Hemagglutinin is the predominant surface glycoprotein and the primary determinant of antigenicity, virulence and zoonotic potential. Mutations leading to changes in the number of HA glycosylation sites are often reported. Such genetic sequencing studies predict at best the disruption or creation of sequons for N-linked glycosylation; they do not reflect actual phenotypic changes in HA structure. Therefore, combined analysis of glycan micro and macro-heterogeneity and bioassays will better define the relationships among glycosylation, viral bioactivity and evolution. We present a study that integrates proteomics, glycomics and glycoproteomics of HA before and after adaptation to innate immune system pressure. We combined this information with glycan array and immune lectin binding data to correlate the phenotypic changes with biological activity. Underprocessed glycoforms predominated at the glycosylation sites found to be involved in viral evolution in response to selection pressures and interactions with innate immune-lectins. To understand the structural basis for site-specific glycan microheterogeneity at these sites, we performed structural modeling and molecular dynamics simulations. We observed that the presence of immature, high-mannose type glycans at a particular site correlated with reduced accessibility to glycan remodeling enzymes. Further, the high mannose glycans at sites implicated in immune lectin recognition were predicted to be capable of forming trimeric interactions with the immune-lectin surfactant protein-D.

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“…The final pFastBac-GP67-H6HA1-His-RhPV-IRES-EGFP construct was transferred to DH10Bac competent cells (Thermo Fisher Scientific) and the recombinant bacmid was produced according to the manufacturer's instructions. Four potential N-linked glycosylation sites on H6HA1-His (Asn-11, Asn-23, Asn-167, and Asn-291) were mutated to alanine through PCR-based site-directed mutagenesis (He et al, 2015).…”

Section: Construction Of the Bi-cistronic Baculovirus Expression Vectormentioning

confidence: 99%

“…HA is a vital antigen, but it is inclined to mutate to escape the attack from the immune system (Van de Sandt et al, 2012). Glycosylation on HA may play a critical role in triggering an immune response (Wang et al, 2009;He et al, 2015). Traditional human inactivated vaccines for influenza are produced by using the specific-pathogen-free (SPF) embryonated egg method, which requires specialized equipment and complex procedures (Nunez et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

A secretary bi-cistronic baculovirus expression system with improved production of the HA1 protein of H6 influenza virus in insect cells and Spodoptera litura larvae

Hsieh

et al. 2018

Journal of Immunological Methods

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A bi-cistronic baculovirus expression vector was constructed to facilitate the expression, detection, and isolation of the hemagglutinin (HA) fragment HA1 of H6N1 avian influenza virus (AIV) in an insect and a culture of its cells. In this construct, the GP67sp signal peptide promoted the secretion of the recombinant protein into the culture medium, and improved protein expression and purification. Enhanced green fluorescent protein, co-expressed through an internal ribosome entry site, served as a visible reporter for protein expression detection. The hemolymph of Spodoptera litura larvae infected with the bi-cistronic baculovirus was collected for the purification of the recombinant HA1, which was found to be glycosylated, and monomeric and trimeric forms of the recombinant HA1 were identified. Proteins expressed in both the cell culture and larvae served as effective subunit vaccines for the production of antiserum against HA. The antiserum recognized the H6 subtype of AIV but not the H5 subtype.

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Glycosylation at hemagglutinin Asn-167 protects the H6N1 avian influenza virus from tryptic cleavage at Arg-201 and maintains the viral infectivity

Cited by 7 publications

References 35 publications

Structural and Functional Studies of Influenza Virus A/H6 Hemagglutinin

Structural and Functional Studies of Influenza Virus A/H6 Hemagglutinin

Integrated Omics and Computational Glycobiology Reveal Structural Basis for Influenza A Virus Glycan Microheterogeneity and Host Interactions

A secretary bi-cistronic baculovirus expression system with improved production of the HA1 protein of H6 influenza virus in insect cells and Spodoptera litura larvae

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