Envelope protein
E protein is a type 1 transmembrane protein able to form pentamers creating pores with ion transport activity. E protein localizes mainly in the ER and Golgi apparatus, where it participates in assembly, budding and intracellular trafficking of newly formed virions. E protein is involved in tight junctions disruption in the lungs to reach the alveolar wall and develop into a systemic infection, triggering an overexpression of inflammatory cytokines and lymphopenia. Four variations in SARS-CoV-2 E protein relative to SARS-CoV-1 were verified, all located in the C-terminal to the cytoplasmic face, thereby potentially modulating E protein interaction with other proteins.
Narrative
The envelope (E) protein is one of the four structural proteins of coronaviruses. The other structural proteins are the membrane (M), nucleocapsid (N) and spike (S) proteins. The E protein is a type 1 transmembrane protein that is able to form pentamers by associating with other E proteins (Li et al. 2014). This pentamer forms a membrane pore that is able to transport ions (Li et al. 2014). The pore structure is called viroporin and it is present in many common viruses. The ion channel activity can be inhibited in SARS-CoV-1 E protein by the drug HMA (Li et al. 2014). Only a few E proteins are present in each viral particle, but it is highly expressed in the host cells (Nieto-Torres et al. 2011; Vennema et al. 1996). The E protein has been proposed to initiate membrane curvature together with the M protein via the interaction between their C-terminal domains, but this mechanism remains largely unknown (Schoeman and Fielding 2019; Lim and Liu 2001). The M protein alone seems to not trigger a proper membrane curvature for virion production. The M protein alone can produce virions, but the absence of the E protein cripples virion production, morphology, and plaque shape (Fischer et al. 1998; L. Kuo and Masters 2003; Vennema et al. 1996; Lim and Liu 2001). The E protein localizes mainly in the ER and Golgi apparatus, where it participates in assembly, budding, and intracellular trafficking of newly formed virions (Vennema et al. 1996; Lim and Liu 2001).
The E protein has been shown to interact with other viral proteins. Tandem affinity purification assays established interaction between the S and the E proteins, but the mechanism of how this happens was not pursued further (Alvarez et al. 2010). This study also shows that the E protein interacts with the nsp3 and they suggest that nsp3 mediates E protein ubiquitination. Besides, the E protein co-immunoprecipitates with the N protein, but the function of this interaction remains unclear (Maeda, Maeda, and Makino 1999). Furthermore, a yeast two-hybrid system and an in vitro pull down assay showed interaction between E proteins and ORF7a, but its importance is yet to be identified (Pan et al. 2008; Fielding et al. 2006). On the other hand, the E protein PDZ-Binding Motif (PBM) interacts with PDZ domains of host proteins. For instance, the interaction of the E protein and a protein associated with Caenorhabditis elegans, lin-7 protein 1 (PALS1), a PDZ-containing protein, disrupts tight junctions in the lungs to reach the alveolar wall and develops into a systemic infection (Teoh et al. 2010). Further, the interaction with syntenin caused it to concentrate in the cytoplasm, triggering an overexpression of inflammatory cytokines, which activates an exaggerated immune response, resulting in lung tissue damage, edema accumulation, and leading to acute respiratory distress syndrome (Jimenez-Guardeño et al. 2014). Interaction between the E protein and the Bcl-xL protein caused lymphopenia (Y. Yang et al. 2005).
Structural analysis and comparison with SARS-CoV-1 envelope protein – Several functionally important residues in E protein have been identified in SARS-CoV-1. For instance, there is a conserved proline residue in the C-terminal in the β-coil-β motif, that if mutated changes localization of the E protein from the Golgi complex to the plasma membrane (Cohen, Lin, and Machamer 2011; Li et al. 2014). Besides, the mutations Asn15Ala and Val25Phe inhibit ion channel activity of SARS-CoV-1 E protein. After several passages through cell cultures, this function is restored by the addition of new mutations, suggesting that the E protein confers a selective advantage to the virus (Nieto-Torres et al. 2014). It is also important to note that the C-terminal of the E protein interacts with the C-terminal of the M protein in the cytoplasmic side, and this is required to virus envelope formation (Lim and Liu 2001). As being highly conserved relative to SARS-CoV-1 (96% identical), this information is likely applicable to SARS-CoV-2 E protein.
SARS-CoV-2 E protein has a predicted short 11 aa N-terminal tail, 25 aa transmembrane region and 37 aa C-terminal cytoplasmic region, including PBM (DLLV). Four variations relative to SARS-CoV-1 E protein were verified, all located in the C-terminal end, including in one conservative substitution, two non-conservative substitutions (Val56Phe, Glu69Arg), and a deletion. Similar to SARS-CoV-1, SARS-CoV-2 E protein is likely to assemble as a pentamer to form a viroporin, and the three C-terminal substitutions are likely exposed in the pentamer to the cytoplasm, thereby they could be involved in modulating E protein interaction with other proteins.
