Plasma membrane-porating domain in poliovirus 2B protein. A short peptide mimics viroporin activity.
Picornavirus 2B, a non-structural protein required for effective viral replication, has been implicated in cell membrane permeabilization during the late phases of infection. Here, we have approached the molecular mechanism of this process by assessing the pore-forming activity of an overlapping peptide library that spanned the complete 2B sequence. At non-cytopathic concentrations, only the P3 peptide, spanning 2B residues 35-55, effectively assembled hydrophilic pores that allowed diffusion of low molecular mass solutes across the cell plasma membrane (IC(50) approximately 4x10(-7) M) and boundary liposome bilayers (starting at peptide to lipid molar ratios>1:10(4)). Circular dichroism data were consistent with its capacity to fold as a helix in a membrane-like environment. Furthermore, addition of this peptide to a sealed plasma-membrane model, consisting of retinal rod outer segments patch-clamped in a whole-cell configuration, induced ion channel activity within seconds at concentrations as low as 10(-8) M. Thus, we have established a "one-helix" 2B version that possesses the intrinsic pore-forming activity required to directly and effectively permeabilize the cell plasma membrane. We conclude that 2B viroporin can be classified as a genuine pore-forming toxin of viral origin, which is produced intracellularly at certain times post infection.
Madan
Vanesa
V
Centro de Biología Molecular (CSIC-UAM), Universidad Autónoma de Madrid, Canto Blanco, 28049 Madrid, Spain.
Sánchez-Martínez
Silvia
S
Vedovato
Natascia
N
Rispoli
Giorgio
G
Carrasco
Luis
L
Nieva
José L
JL
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D002463:Cell Membrane Permeability; D008081:Liposomes / Q000737:chemistry; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000502:physiology*
C101758:2B protein, poliovirus; D008081:Liposomes; D019151:Peptide Library; D010455:Peptides; D017361:Viral Nonstructural Proteins