Biology subjects

Unione, L.

Publications and source records attributed to Unione, L..

4 recordsLinked to original sources

bioRxiv2024

Epistasis in the receptor binding domain of contemporary H3N2 viruses that reverted to bind sialylated diLacNAc repeats

Since the introduction of H3N2 influenza A viruses in the human population, these viruses have continuously evolved to escape human immunity, with mutations occurring in and around the receptor binding site. This process, called antigenic drift, recently resulted in viruses that recognize elongated glycans that are not abundantly displayed in the human respiratory tract. Such receptor specificities hampered our ability to pick and propagate vaccine strains. Using ELISA, glycan array, tissue staining, flow cytometry, and hemagglutinin assays, this study revealed that the most recent H3N2 viruses have expanded receptor specificity by regaining effective recognition to shorter glycans. In recent H3 strains, Y159 and T160 are responsible for restricted binding to elongated glycans; in contemporary strains, however, Y159N and T160I dominate with a consequent loss of strength in receptor binding. Yet, effective receptor interaction is rescued by a remote mutation in the 190-helix, Y195F. The results demonstrate epistasis of critical residues in three of the four structural elements composing the HA receptor-binding site (the 130-loop, 150-loop, and 190-helix), which synergistically contribute to shape receptor binding specificity. Interestingly, a positive correlation exists between binding to an asymmetrical N-glycan containing an 2,6 sialylated tri-LacNAc arm and binding to human and ferret respiratory tract tissues. Together, these results elucidate the epistatic nature of receptor binding specificity during influenza A virus H3N2 evolution.

microbiology↗

bioRxiv2023

Probing Altered Receptor Specificities of Antigenically Drifting Human H3N2 Viruses by Chemoenzymatic Synthesis, NMR and Modeling

Prototypic receptors for human influenza viruses are cell surface N-glycans carrying 2,6-linked sialosides. Under immune pressure, A/H3N2 influenza viruses have emerged with altered receptor specificities that appear to recognize 2,6-linked sialosides presented on extended N-acetyl-lactosamine (LacNAc) moieties. Here, molecular recognition features of such drifted hemagglutinins (HAs) are examined by chemoenzymatic synthesis of complex N-glycans having 13C-labeled monosaccharides at strategic positions. The labeled glycans were employed in 2D STD-1H and 13C-HSQC NMR experiments to pinpoint which monosaccharides of the extended LacNAc chain engage with evolutionarily distinct HAs. The NMR data in combination with computational and mutagenesis studies demonstrate that mutations distal to the receptor binding domain of recent HAs have created an extended binding site that can directly interact with the extended LacNAc chain. A fluorine containing sialyl-LacNAc derivative is used as NMR probe to derive relative binding affinities and confirmed the contribution of the extended LacNAc chain for binding.

biochemistry↗

bioRxiv2022

The SARS-CoV-2 spike N-terminal domain engages 9-O-acetylated α2-8-linked sialic acids

SARS-CoV-2 viruses engage ACE2 as a functional receptor with their spike protein. The S1 domain of the spike protein contains a C-terminal receptor-binding domain (RBD) and an N-terminal domain (NTD). The NTD of other coronaviruses includes a glycan-binding cleft. However, for the SARS-CoV-2 NTD protein-glycan binding was only observed weakly for sialic acids with highly sensitive methods. Amino acid changes in the NTD of Variants of Concern (VoC) shows antigenic pressure, which can be an indication of NTD-mediated receptor binding. Trimeric NTD proteins of SARS-CoV-2, Alpha, Beta, Delta, and Omicron did not reveal a receptor binding capability. Unexpectedly, the SARS-CoV-2 Beta subvariant strain (501Y.V2-1) NTD binding to Vero E6 cells was sensitive to sialidase pretreatment. Glycan microarray analyses identified a putative 9-O-acetylated sialic acid as a ligand, which was confirmed by catch-and-release ESI-MS, STD-NMR analyses, and a graphene-based electrochemical sensor. The Beta (501Y.V2-1) variant attained an enhanced glycan binding modality in the NTD with specificity towards 9-O-acetylated structures, suggesting a dual-receptor functionality of the SARS-CoV-2 S1 domain, which was quickly selected against. These results indicate that SARS-CoV-2 can probe additional evolutionary space, allowing binding to glycan receptors on the surface of target cells. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=76 HEIGHT=200 SRC="FIGDIR/small/507904v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1f98a70org.highwire.dtl.DTLVardef@1efc119org.highwire.dtl.DTLVardef@16e8bc6org.highwire.dtl.DTLVardef@9a74e6_HPS_FORMAT_FIGEXP M_FIG C_FIG SynopsisCoronaviruses utilize their N-terminal domain (NTD) for initial reversible low-affinity interaction to (sialylated) glycans. This initial low-affinity/high-avidity engagement enables viral surfing on the target membrane, potentially followed by a stronger secondary receptor interaction. Several coronaviruses, such as HKU1 and OC43, possess a hemagglutinin-esterase for viral release after sialic acid interaction, thus allowing viral dissemination. Other coronaviruses, such as MERS-CoV, do not possess a hemagglutinin-esterase, but interact reversibly to sialic acids allowing for viral surfing and dissemination. The early 501Y.V2-1 subvariant of the Beta SARS-CoV-2 Variant of Concern has attained a receptor-binding functionality towards 9-O-acetylated sialic acid using its NTD. This binding functionality was selected against rapidly, most likely due to poor dissemination. Ablation of sialic acid binding in more recent SARS-CoV-2 Variants of Concern suggests a fine balance of sialic acid interaction of SARS-CoV-2 is required for infection and/or transmission.

molecular biology↗

bioRxiv2020

Glycan remodeled erythrocytes facilitate antigenic characterizationof recent A/H3N2 influenza viruses

During circulation in humans and natural selection to escape antibody recognition for decades, A/H3N2 influenza viruses emerged with altered receptor specificities. These viruses lost the ability to agglutinate erythrocytes critical for antigenic characterization and give low yields and acquire adaptive mutations when cultured in eggs and cells, contributing to recent vaccine challenges. We examined receptor specificities of A/H3N2 viruses, revealing that recent viruses compensated for decreased binding of the prototypic human receptor by recognizing 2,6-sialosides on extended LacNAc moieties. Erythrocyte glycomics showed an absence of extended glycans, providing a rationale for lack of agglutination by recent A/H3N2 viruses. A glycan remodeling approach installed functional receptors on erythrocytes, allowing antigenic characterization of recent A/H3N2 viruses and confirming the cocirculation of several antigenically different viruses in humans. Computational studies of HAs in complex with a sialoside having an extended LacNAc moiety revealed that mutations distal to the RBD reoriented the Y159 side chain, resulting in an extended receptor binding site.

biochemistry↗