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J Mol Biol. 2019 Aug 9;431(17):3261-3274. doi: 10.1016/j.jmb.2019.05.044. Epub 2019 Jun 4.

The Arginines in the N-Terminus of the Porcine Circovirus 2 Virus-like Particles Are Responsible for Disrupting the Membranes at Neutral and Acidic pH.

Author information

1
Department of Chemistry and Biochemistry, The City College of New York, New York, NY 10031, USA.
2
Department of Chemistry and Biochemistry, The City College of New York, New York, NY 10031, USA; Graduate Program in Biochemistry, The Graduate Center of the City University of New York, New York, NY 10016, USA.
3
Department of Chemistry and Biochemistry, The City College of New York, New York, NY 10031, USA; Graduate Program in Biochemistry, The Graduate Center of the City University of New York, New York, NY 10016, USA. Electronic address: rkhayat@ccny.cuny.edu.

Abstract

Non-enveloped viruses that are endocytosed employ numerous mechanisms to disrupt endosomal membranes for escape into the cellular cytoplasm. These include the use of amphipathic helices or sheets, hydrophobic loops, myristoylated peptides, and proteins with phospholipase activity. Some mechanisms result in immediate deterioration of the endosome, while others form pores in the membrane causing osmolysis to disrupt the endosome and allow viral escape. We describe an additional mechanism by a non-enveloped virus to disrupt endosomal membranes. Porcine circovirus 2 (PCV2) possesses a 41-amino acid arginine-rich motif (ARM) at the N-terminus of its capsid protein that appears to be in the interior of the virus-like particle (VLP). Using in vitro membrane disruption assays, we demonstrate that PCV2 VLP, unassembled capsid, and ARM peptide possess the ability to disrupt endosomal-like membranes, whereas VLP lacking the ARM sequence does not possess this capability. Membrane disruption by VLP is insensitive to pH, but unassembled capsid protein and ARM peptide exhibit diminished activity at low pH. Our liposome disruption assays, circular dichroism, and intrinsic tryptophan fluorescence assays allow us to propose a model for PCV2-endosomal membrane interaction wherein the ARM peptide externalizes from the capsid, its C-terminus (amino acids 28-40) anchors into the membrane, and the arginine-rich N-terminus (amino acids 1-27) drives membrane disruption. To our knowledge, this is the first example of a non-enveloped virus using the arginines of an ARM to disrupt membranes. Also, this is the first example of such study for the Circoviridae family of viruses.

KEYWORDS:

arginine-rich motif; capsid protein; endosomal escape; liposome disruption; porcine circovirus

PMID:
31173778
PMCID:
PMC6697213
[Available on 2020-08-09]
DOI:
10.1016/j.jmb.2019.05.044

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