Middle East Respiratory Syndrome coronavirus (MERS-CoV) causes severe pulmonary infection, with â¼35 % mortality. Spike glycoprotein (S) of MERS-CoV is a key target for vaccines and therapeutics because S mediates viral entry and membrane-fusion to host cells. Here, four different S subunit proteins, receptor-binding domain (RBD; 358-606 aa), S1 (1-751 aa), S2 (752-1296 aa), and S&#916;TM (1-1296 aa), were generated using the baculoviral system and immunized in mice to develop neutralizing antibodies. We developed 77 hybridomas and selected five neutralizing mAbs by immunization with S&#916;TM against MERS-CoV EMC/2012 strain S-pseudotyped lentivirus. However, all five monoclonal antibodies (mAb) did not neutralize the pseudotyped V534A mutation. Additionally, one mAb RBD-14F8 did not show neutralizing activity against pseudoviruses with amino acid substitution of L506â¯F or D509â¯G (England1 strain, EMC/2012 L506â¯F, and EMC/2012 D509â¯G), and RBD-43E4 mAb could not neutralize the pseudotyped I529â¯T mutation, while three other neutralizing mAbs showed broad neutralizing activity. This implies that the mutation in residue 506-509, 529, and 534 of S is critical to generate neutralization escape variants of MERS-CoV. Interestingly, all five neutralizing mAbs have binding affinity to RBD, although most mAbs generated by RBD did not have neutralizing activity. Additionally, chimeric antibodies of RBD-14F8 and RBD-43E4 with human Fc and light chain showed neutralizing effect against wild type MERS-CoV KOR/KNIH/002, similar to the original mouse mAbs. Thus, our mAbs can be utilized for the identification of specific mutations of MERS-CoV.
Characterization of novel monoclonal antibodies against MERS-coronavirus spike protein