FIG. 1.

(A) Spike constructs. MHV-A59 S was expressed from the plasmid pTUMS (32), and the FIPV strain 79-1146 S protein was expressed from pFIPVE2, which was made as follows. A 3′-terminal S fragment was prepared by ligating the XbaI-SalI fragment from pB1 (4) into pUC18, cutting with AccI and SalI, and religating after filling in with Klenow polymerase. From the resulting plasmid p3d, the XbaI-SalI fragment was isolated and used. A middle piece was prepared by isolating the PstI-XbaI fragment from pB1. This fragment and the 3′ XbaI-SalI fragment were ligated into p1A (4), which had been digested with PstI and SalI to give pFIPVE2. Chimeric protein FMS was expressed from pTFMS, which was constructed as follows. Plasmid p3d was digested with HindIII, filled in with Klenow enzyme, and ligated with BglII linkers, resulting in p3dHrB. After the plasmid was cut with StyI and BglII, an MHV S gene fragment was ligated into it; the fragment was prepared by digesting the S gene, obtained as a BamHI fragment from pDGE2 (31), with StyI and taking the small fragment. The resulting p3FM vector was cut with PstI and SalI; into it were ligated the XbaI-SalI fragment from p3d and the PstI-XbaI fragment from pB1. The chimeric gene was finally recloned as a BamHI fragment into pTUG3, resulting in pTFMS. Chimeric protein MFS was expressed from pTMFS, which was prepared starting with p3dHrB. This plasmid was cut with StyI and BamHI, and a BamHI-StyI fragment obtained from the MHV S BamHI gene described above was ligated into it. The chimeric S gene was recloned as a BamHI-SalI fragment into pTUG3 cut with the same enzymes. TM, transmembrane domain; ecto, ectodomain; endo, endodomain. (B) Carboxy-terminal sequences of the MHV-A59 and FIPV spike proteins. The 67 terminal residues of each protein are compared. The arrow indicates the junction point in the chimeric S constructs.