Identification of amino acid residues in BK virus VP1 that are critical for viability and growth.
BK virus (BKV) is a ubiquitous pathogen that establishes a persistent infection in the urinary tract of 80% of the human population. Like other polyomaviruses, the major capsid protein of BKV, virion protein 1 (VP1), is critical for host cell receptor recognition and for proper virion assembly. BKV uses a carbohydrate complex containing alpha(2,3)-linked sialic acid attached to glycoprotein and glycolipid motifs as a cellular receptor. To determine the amino acids important for BKV binding to the sialic acid portion of the complex, we generated a series of 17 point mutations in VP1 and scored them for viral growth. The first set of mutants behaved identically to wild-type virus, suggesting that these amino acids were not critical for virus propagation. Another group of VP1 mutants rendered the virus nonviable. These mutations failed to protect viral DNA from DNase I digestion, indicating a role for these domains in capsid assembly and/or packaging of DNA. A third group of VP1 mutations packaged DNA similarly to the wild type but failed to propagate. The initial burst size of these mutations was similar to that of the wild type, indicating that there is no defect in the lytic release of the mutated virions. Binding experiments revealed that a subset of the BKV mutants were unable to attach to their host cells. These motifs are likely important for sialic acid recognition. We next mapped these mutations onto a model of BKV VP1 to provide atomic insight into the role of these sites in the binding of sialic acid to VP1.
Dugan
Aisling S
AS
Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, 70 Ship Street, Providence, RI 02903, USA.
Gasparovic
Megan L
ML
Tsomaia
Natia
N
Mierke
Dale F
DF
O'Hara
Bethany A
BA
Manley
Kate
K
Atwood
Walter J
WJ
D000595:Amino Acid Sequence; D000818:Animals; D001739:BK Virus / Q000378:metabolism*; D002213:Capsid / Q000737:chemistry; D036022:Capsid Proteins / Q000737:chemistry / Q000502:physiology*; D002470:Cell Survival; D002522:Chlorocebus aethiops; D006023:Glycoproteins / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019158:N-Acetylneuraminic Acid / Q000737:chemistry; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D014709:Vero Cells
D036022:Capsid Proteins; D006023:Glycoproteins; C062481:VP1 protein, polyomavirus; D019158:N-Acetylneuraminic Acid
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