Identification of a conserved linear B-cell epitope in the M protein of porcine epidemic diarrhea virus

BACKGROUND: The major structural protein of coronaviruses, the membrane (M) protein, can elicit the formation of protective antibodies, but little information is available about the M protein of porcine epidemic diarrhea virus (PEDV). Identification of epitopes on the PEDV M protein will be helpful in the elucidation of the antigenic properties of this protein. RESULTS: One hybridoma cell line secreting anti-M protein monoclonal antibody (McAb) was generated and designated 4D4. To map the epitopes on the PEDV M protein, a total of 17 partially overlapping fragments covering the C-terminus of M protein were expressed as fusion proteins with a 6×His tag or a GST tag. A linear motif, (193)TGWAFYVR(200), was identified by enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis using McAb 4D4. The motif (195)WAFYVR(200) was the minimal requirement for reactivity, as demonstrated by removing amino acids individually from both ends of the motif (193)TGWAFYVR(200). The result of WB analysis showed that the 4D4-defined epitope could be recognized by PEDV-positive serum, but not transmissible gastroenteritis virus (TGEV)-positive serum. Furthermore, this epitope was highly conserved among different PEDV strains, as shown by alignment and comparison of sequences. CONCLUSION: A McAb, 4D4, directed against the M protein of PEDV, was obtained, and the 4D4-defined minimal epitope sequence was (195)WAFYVR(200). The McAb could serve as a candidate for development of a McAb-based antigen capture ELISA for detection of PEDV. The epitope identified provides a basis for the development of epitope-based differential diagnostic techniques and may be useful in the design of epitope-based vaccines.