Proteolytic cleavage of the E2 glycoprotein of murine coronavirus: activation of cell-fusing activity of virions by trypsin and separation of two different 90K cleavage fragments

Abstract

In the murine coronavirus mouse hepatitis virus, a single glycoprotein, E2, is required both for attachment to cells and for cell fusion. Cell fusion induced by infection with mouse hepatitis virus strain A59 was inhibited by the addition of monospecific anti-E2 antibody after virus adsorption and penetration. Adsorption of concentrated coronavirions to uninfected cells did not cause cell fusiOn in the presence of cycloheximide. Thus, cell fusion was induced by E2 on the plasma membrane of infected 17 Cl I cells but not by E2 on virions grown in these cells. Trypsin treatment of virions purified from 17 Cl 1 cells quantitatively cleaved 180K E2 to 90K E2 and activated cell-fusing activity of the virions. This proteolytic cleavage yielded two different 90K species which were separable by sodium dodecyl sulfate-hydroxyapatite chromatography. One of the trypsin cleavage products, 90A, was acylated and may be associated with the lipid bilayer. The other, 90B, was not acylated and yielded different peptides than did 90A upon limited digestion with thermolysin or staphylococcal V8 protease. Thus, the cell-fusing activity of a coronavirus required proteolytic cleavage of the E2 glycoprotein, either by the addition of a protease to virions or by cellular proteases acting on E2, which was transported to the plasma membrane during virus maturation. There is a striking functional similarity between the E2 glycoprotein of coronavirus, which is a positive-strand RNA virus, and the hemagglutinin glycoprotein of negative-strand orthomyxoviruses, in that a single glycoprotein has both attachment and protease-activated cell-fusing activities.

Lactogenic immunity and vaccines for porcine epidemic diarrhea virus (PEDV): Historical and current concepts

“…The S protein forms the characteristic protruding spikes that are situated on the outside of the nucleocapsid and consists of two domains, S1 and S2. The S1 domain is associated with binding to host cellular receptors while the S2 mediates viral fusion and entry into the cell (Bosch et al, 2003;Sturman et al, 1985;Wicht et al, 2014). For TGEV, the S1 domain is a potent inducer of virus neutralizing antibodies, which are important for piglet protection.…”

Structures and Polymorphic Interactions of Two Heptad-Repeat Regions of the SARS Virus S2 Protein

“…Coronavirus S proteins are synthesized as single-chain precursors that oligomerize in the endoplasmic reticulum and are processed through the Golgi, eventually forming long, petal-shaped spikes that protrude from the virion surface (Lai and Holmes, 2001). In some coronaviruses, S is posttranslationally cleaved into two chains, known as S1 (the receptor binding protein) and S2 (the transmembrane fusion protein) (Frana et al, 1985;Sturman et al, 1985). Coronavirus S2 is functionally and structurally related to a large group of so-called class I viral fusion proteins, including those of orthomyxoviruses such as influenza virus, paramyxoviruses such as SV5, retroviruses such as HIV, and filoviruses such as Ebola virus (Earp et al, 2005;Eckert and Kim, 2001;Harrison, 2005;Weissenhorn et al, 1999).…”

Analysis of protein expression changes of the Vero E6 cells infected with classic PEDV strain CV777 by using quantitative proteomic technique

“…Cystatin-C has been shown to reduce the replication of certain viruses, including the poliovirus, rhinovirus, and human coronaviruses OC43 and 229E (Korant et al, 1986;Collins and Grubb, 1991). The cleavage of S protein has been shown to be essential for the induction of cell-to-cell fusion and coronavirus entry into cells (Sturman et al, 1985). Shirato et al (2011) reported the transmembrane type II serine protease 2 enhanced infection of PEDV in Vero cells by increasing virus release.…”