Identification of a Membrane Binding Peptide in the Envelope Protein of MHV Coronavirus

Coronaviruses (CoVs) are enveloped, positive sense, single strand RNA viruses that cause respiratory, intestinal and neurological diseases in mammals and birds. Following replication, CoVs assemble on intracellular membranes including the endoplasmic reticulum Golgi intermediate compartment (ERGIC) where the envelope protein (E) functions in virus assembly and release. In consequence, E potentially contains membrane-modifying peptides. To search for such peptides, the E coding sequence of Mouse Hepatitis Virus (MHV) was inspected for its amino acid conservation, proximity to the membrane and/or predicted amphipathic helices. Peptides identified in silico were synthesized and tested for membrane-modifying activity in the presence of giant unilamellar vesicles (GUVs) consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), sphingomyelin and cholesterol. To confirm the presence of membrane binding peptides identified in the context of a full-length E protein, the wild type and a number of mutants in the putative membrane binding peptide were expressed in Lenti-X-293T mammalian and insect cells, and the distribution of E antigen within the expressing cell was assessed. Our data identify a role for the post-transmembrane region of MHV E in membrane binding.