The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors

María Ríos Carrasco; Ting-Hui Lin; Xueyong Zhu; Alba Gabarroca García; Elif Uslu; Ruonan Liang; Cindy M Spruit; Mathilde Richard; Geert-Jan Boons; Ian A Wilson; Robert P de Vries

doi:10.1073/pnas.2419800122

The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors

María Ríos Carrasco
, Ting-Hui Lin
, Xueyong Zhu
, Alba Gabarroca García
, Elif Uslu
, Ruonan Liang
, Cindy M Spruit
, Mathilde Richard
, Geert-Jan Boons
, Ian A Wilson
, Robert P de Vries

Scripps Research Institute
Utrecht University
Erasmus University Medical Center Rotterdam

Research output: Contribution to journal › Article › Academic › peer-review

Abstract

H5Nx viruses continue to wreak havoc in avian and mammalian species worldwide. The virus distinguishes itself by the ability to replicate to high titers and transmit efficiently in a wide variety of hosts in diverse climatic environments. Fortunately, transmission to and between humans is scarce. Yet, if such an event were to occur, it could spark a pandemic as humans are immunologically naïve to H5 viruses. A significant determinant of transmission to and between humans is the ability of the influenza A virus hemagglutinin (HA) protein to shift from an avian-type to a human-type receptor specificity. Here, we demonstrate that a 2016 2.3.4.4e virus HA can convert to human-type receptor binding via a single Q226L mutation, in contrast to a cleavage-modified 2016 2.3.4.4b virus HA. Using glycan arrays, X-ray structural analyses, tissue- and direct glycan binding, we show that L133a Δ and 227Q are vital for this phenotype. Thus, whereas the 2.3.4.4e virus HA only needs a single amino acid mutation, the modified 2016 2.3.4.4b HA was not easily converted to human-type receptor specificity.

Original language	English
Article number	e2419800122
Journal	Proceedings of the National Academy of Sciences of the United States of America
Volume	122
Issue number	16
DOIs	https://doi.org/10.1073/pnas.2419800122
Publication status	Published - 15 Apr 2025

Bibliographical note

Publisher Copyright:
Copyright © 2025 the Author(s).

Funding

R.P.d.V. is a recipient of an ERC Starting grant from the European Commission (802780) and is supported by the Mizutani Foundation for Glycoscience. This research was made possible by funding from International coordination of research on infectious diseases, an European Research Area Network cofunded under the European Union\u2019s Horizon 2020 research and innovation programme (https://ec.europa.eu/programmes/horizon2020/en), under Grant Agreement no. 862605 (Flu-Switch) to R.P.d.V. R.P.d.V and M.R.C are supported by an NWO-M2 (OCENW.M20.106). The glycan array setup was supported by the Netherlands Organization for Scientific Research (NWO, TOP-PUNT 718.015.003 to G.-J.B.). This structural biology research at Scripps Research was partially funded by NIH National Institute of Allergy and Infectious Diseases Centers of Excellence for Influenza Research and Response contract 75N93021C00015/PENN CEIRR (I.A.W.). X-ray diffraction datasets were collected at the Stanford Synchrotron Radiation Lightsource (SSRL) beamline 12-1. Use of the SSRL, Stanford Linear Accelerator Center National Accelerator Laboratory, is supported by the US Department of Energy (DOE), Office of Science, Office of Basic Energy Sciences under Contract No. DE-AC02-76SF00515. The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research and by the NIH, National Institute of General Medical Sciences (P30GM133894). The contents of this publication are solely the responsibility of the authors and do not necessarily represent the official views of National Institute of General Medical Sciences or NIH. We gratefully acknowledge all data contributors, i.e., the authors and their originating laboratories responsible for obtaining the specimens and their submitting laboratories for generating the genetic sequence and metadata and sharing via the Global Initiative on Sharing All Influenza Data Initiative, on which part of this research is based.

Funders	Funder number
Stanford Linear Accelerator Center National Accelerator Laboratory
Biological and Environmental Research
Mizutani Foundation for Glycoscience
SSRL
National Institutes of Health
U.S. Department of Energy
Office of Science
NIH National Institute of Allergy and Infectious Diseases Centers of Excellence for Influenza Research and Response
European Commission	802780
National Institute of General Medical Sciences	P30GM133894
Horizon 2020 Framework Programme	862605
Basic Energy Sciences	DE-AC02-76SF00515
Nederlandse Organisatie voor Wetenschappelijk Onderzoek	TOP-PUNT 718.015.003
NWO-M2	OCENW.M20.106

Keywords

Animals
HEK293 Cells
Hemagglutinin Glycoproteins, Influenza Virus/genetics
Humans
Influenza A virus/genetics
Influenza in Birds/virology
Influenza, Human/virology
Mutation
Protein Binding
Receptors, Virus/metabolism
hemagglutinin
sialic acid
glycan array
influenza A virus

Access to Document

10.1073/pnas.2419800122

ríos-carrasco-et-al-the-q226l-mutation-can-convert-a-highly-pathogenic-h5-2-3-4-4e-virus-to-bind-human-type-receptorsFinal published version, 2.11 MBLicence: Taverne

Cite this

Ríos Carrasco, M., Lin, T.-H., Zhu, X., Gabarroca García, A., Uslu, E., Liang, R., Spruit, C. M., Richard, M., Boons, G.-J., Wilson, I. A., & de Vries, R. P. (2025). The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors. Proceedings of the National Academy of Sciences of the United States of America, 122(16), Article e2419800122. https://doi.org/10.1073/pnas.2419800122

@article{8a32cd3793bd48dabb5eab4f33986711,

title = "The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors",

abstract = "H5Nx viruses continue to wreak havoc in avian and mammalian species worldwide. The virus distinguishes itself by the ability to replicate to high titers and transmit efficiently in a wide variety of hosts in diverse climatic environments. Fortunately, transmission to and between humans is scarce. Yet, if such an event were to occur, it could spark a pandemic as humans are immunologically na{\"i}ve to H5 viruses. A significant determinant of transmission to and between humans is the ability of the influenza A virus hemagglutinin (HA) protein to shift from an avian-type to a human-type receptor specificity. Here, we demonstrate that a 2016 2.3.4.4e virus HA can convert to human-type receptor binding via a single Q226L mutation, in contrast to a cleavage-modified 2016 2.3.4.4b virus HA. Using glycan arrays, X-ray structural analyses, tissue- and direct glycan binding, we show that L133a Δ and 227Q are vital for this phenotype. Thus, whereas the 2.3.4.4e virus HA only needs a single amino acid mutation, the modified 2016 2.3.4.4b HA was not easily converted to human-type receptor specificity.",

keywords = "Animals, HEK293 Cells, Hemagglutinin Glycoproteins, Influenza Virus/genetics, Humans, Influenza A virus/genetics, Influenza in Birds/virology, Influenza, Human/virology, Mutation, Protein Binding, Receptors, Virus/metabolism, hemagglutinin, sialic acid, glycan array, influenza A virus",

author = "\{R{\'i}os Carrasco\}, Mar{\'i}a and Ting-Hui Lin and Xueyong Zhu and \{Gabarroca Garc{\'i}a\}, Alba and Elif Uslu and Ruonan Liang and Spruit, \{Cindy M\} and Mathilde Richard and Geert-Jan Boons and Wilson, \{Ian A\} and \{de Vries\}, \{Robert P\}",

note = "Publisher Copyright: Copyright {\textcopyright} 2025 the Author(s).",

year = "2025",

month = apr,

day = "15",

doi = "10.1073/pnas.2419800122",

language = "English",

volume = "122",

journal = "Proceedings of the National Academy of Sciences of the United States of America",

issn = "0027-8424",

publisher = "National Academy of Sciences",

number = "16",

}

Ríos Carrasco, M, Lin, T-H, Zhu, X, Gabarroca García, A, Uslu, E , Liang, R, Spruit, CM, Richard, M, Boons, G-J, Wilson, IA & de Vries, RP 2025, 'The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors', Proceedings of the National Academy of Sciences of the United States of America, vol. 122, no. 16, e2419800122. https://doi.org/10.1073/pnas.2419800122

TY - JOUR

T1 - The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors

AU - Ríos Carrasco, María

AU - Lin, Ting-Hui

AU - Zhu, Xueyong

AU - Gabarroca García, Alba

AU - Uslu, Elif

AU - Liang, Ruonan

AU - Spruit, Cindy M

AU - Richard, Mathilde

AU - Boons, Geert-Jan

AU - Wilson, Ian A

AU - de Vries, Robert P

PY - 2025/4/15

Y1 - 2025/4/15

N2 - H5Nx viruses continue to wreak havoc in avian and mammalian species worldwide. The virus distinguishes itself by the ability to replicate to high titers and transmit efficiently in a wide variety of hosts in diverse climatic environments. Fortunately, transmission to and between humans is scarce. Yet, if such an event were to occur, it could spark a pandemic as humans are immunologically naïve to H5 viruses. A significant determinant of transmission to and between humans is the ability of the influenza A virus hemagglutinin (HA) protein to shift from an avian-type to a human-type receptor specificity. Here, we demonstrate that a 2016 2.3.4.4e virus HA can convert to human-type receptor binding via a single Q226L mutation, in contrast to a cleavage-modified 2016 2.3.4.4b virus HA. Using glycan arrays, X-ray structural analyses, tissue- and direct glycan binding, we show that L133a Δ and 227Q are vital for this phenotype. Thus, whereas the 2.3.4.4e virus HA only needs a single amino acid mutation, the modified 2016 2.3.4.4b HA was not easily converted to human-type receptor specificity.

AB - H5Nx viruses continue to wreak havoc in avian and mammalian species worldwide. The virus distinguishes itself by the ability to replicate to high titers and transmit efficiently in a wide variety of hosts in diverse climatic environments. Fortunately, transmission to and between humans is scarce. Yet, if such an event were to occur, it could spark a pandemic as humans are immunologically naïve to H5 viruses. A significant determinant of transmission to and between humans is the ability of the influenza A virus hemagglutinin (HA) protein to shift from an avian-type to a human-type receptor specificity. Here, we demonstrate that a 2016 2.3.4.4e virus HA can convert to human-type receptor binding via a single Q226L mutation, in contrast to a cleavage-modified 2016 2.3.4.4b virus HA. Using glycan arrays, X-ray structural analyses, tissue- and direct glycan binding, we show that L133a Δ and 227Q are vital for this phenotype. Thus, whereas the 2.3.4.4e virus HA only needs a single amino acid mutation, the modified 2016 2.3.4.4b HA was not easily converted to human-type receptor specificity.

KW - Animals

KW - HEK293 Cells

KW - Hemagglutinin Glycoproteins, Influenza Virus/genetics

KW - Humans

KW - Influenza A virus/genetics

KW - Influenza in Birds/virology

KW - Influenza, Human/virology

KW - Mutation

KW - Protein Binding

KW - Receptors, Virus/metabolism

KW - hemagglutinin

KW - sialic acid

KW - glycan array

KW - influenza A virus

UR - https://www.scopus.com/pages/publications/105003448979

U2 - 10.1073/pnas.2419800122

DO - 10.1073/pnas.2419800122

M3 - Article

C2 - 40232794

SN - 0027-8424

VL - 122

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 16

M1 - e2419800122

ER -

The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors

Abstract

Bibliographical note

Funding

Keywords

Access to Document

Other files and links

Fingerprint

The Q226L mutation can convert a highly pathogenic H5 2.3.4.4e virus to bind human-type receptors

Cite this