The receptor-binding domain of influenza virus hemagglutinin produced in Escherichia coli folds into its native, immunogenic structure.
The hemagglutinin (HA) surface glycoprotein promotes influenza virus entry and is the key protective antigen in natural immunity and vaccines. The HA protein is a trimeric envelope glycoprotein consisting of a globular receptor-binding domain (HA-RBD) that is inserted into a membrane fusion-mediating stalk domain. Similar to other class I viral fusion proteins, the fusogenic stalk domain spontaneously refolds into its postfusion conformation when expressed in isolation, consistent with this domain being trapped in a metastable conformation. Using X-ray crystallography, we show that the influenza virus HA-RBD refolds spontaneously into its native, immunogenic structure even when expressed in an unglycosylated form in Escherichia coli. In the 2.10-Å structure of the HA-RBD, the receptor-binding pocket is intact and its conformational epitopes are preserved. Recombinant HA-RBD is immunogenic and protective in ferrets, and the protein also binds with specificity to sera from influenza virus-infected humans. Overall, the data provide a structural basis for the rapid production of influenza vaccines in E. coli. From an evolutionary standpoint, the ability of the HA-RBD to refold spontaneously into its native conformation suggests that influenza virus acquired this domain as an insertion into an ancestral membrane-fusion domain. The insertion of independently folding domains into fusogenic stalk domains may be a common feature of class I viral fusion proteins.
Department of Structural Biology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105, USA.
Rebecca M
RM
DuBois
José Manuel
JM
Aguilar-Yañez
Gonzalo I
GI
Mendoza-Ochoa
Yuriana
Y
Oropeza-Almazán
Stacey
S
Schultz-Cherry
Mario Moisés
MM
Alvarez
Stephen W
SW
White
Charles J
CJ
Russell
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D001665:Binding Sites; D018360:Crystallography, X-Ray; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D005289:Ferrets; D006389:Hemagglutinins, Viral / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D007252:Influenza Vaccines / Q000008:administration & dosage / Q000276:immunology; D008958:Models, Molecular; D009976:Orthomyxoviridae Infections / Q000276:immunology / Q000517:prevention & control; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000914:Antibodies, Viral; D006389:Hemagglutinins, Viral; D007252:Influenza Vaccines; D011994:Recombinant Proteins; C458881:hemagglutinin fusogenic peptide, influenza virus