Human influenza virus hemagglutinin with high sensitivity to proteolytic activation.
To examine the prerequisites for cleavage activation of the hemagglutinin of human influenza viruses, a cDNA clone obtained from strain A/Port Chalmers/1/73 (serotype H3) was subjected to site-directed mutagenesis and expressed in CV-1 cells by using a simian virus 40 vector. The number of basic residues at the cleavage site, which consists of a single arginine with wild-type hemagglutinin, was increased by inserting two, three, or four additional arginines. Like wild-type hemagglutinin, mutants with up to three additional arginines were not cleaved in CV-1 cells, but insertion of four arginines resulted in activation. When the oligosaccharide at asparagine 22 of the HA1 subunit of the hemagglutinin was removed by site-directed mutagenesis of the respective glycosylation site, only three inserted arginines were required to obtain cleavage. Mutants containing a series of four basic residues were also generated by substituting arginine for uncharged amino acids immediately preceding the cleavage site. The observation that these mutants were not cleaved, even when the carbohydrate at asparagine 22 of HA1 was absent, underscores the fact that the basic peptide had to be generated by insertion to obtain cleavage. The data show that the hemagglutinin of a human influenza virus can acquire high cleavability, a property known to be an important determinant for the pathogenicity of avian influenza viruses. Factors important for cleavability are the number of basic residues at the cleavage site, the oligosaccharide at asparagine 22, and the length of the carboxy terminus of HA1.
Ohuchi
R
R
Institut für Virologie, Philipps-Universität Marburg, Germany.
Ohuchi
M
M
Garten
W
W
Klenk
H D
HD
D000595:Amino Acid Sequence; D001120:Arginine / Q000737:chemistry; D001483:Base Sequence; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D010450:Endopeptidases / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D006389:Hemagglutinins, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009980:Influenza A virus / Q000378:metabolism*; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D013329:Structure-Activity Relationship
D006023:Glycoproteins; D006389:Hemagglutinins, Viral; D009841:Oligonucleotides; D001120:Arginine; D010450:Endopeptidases