Epitope mapping.

Seven monoclonal antibodies recognize two antigenic sequences in the supervillin N-terminus. (A) Superimposed representative intensity maps of signals from linear peptide microarrays showing that clones 1E2, 2G3, 4A3 and 5G3 recognize the consensus motif (SA)TRQAHDL, residues 215–223 in hSV1. Peptide overlap makes the requirement for the first two amino acids uncertain. (B) Amplification of the 5A8 intensity map suggests weak binding of clone 5A8 to AERRRQLAEKYGLTL, amino acids 110–124 in hSV1. (C) Recognition on immunoblots of overexpressed, truncated GFP-tagged bovine SV1 (bSV1) proteins in RH30 cell lysates shows that murine hybridoma supernatants (M) 5A8, 4A8 and 5B8 all recognize bSV1-174 and bSV1-127, but not bSV1-101 (D) Alignments showing the sequence conservation of the (SA)TRQAHDL and AERRRQLAEKYGLTL epitopes in mammalian supervillins. Species (reference sequence number): Human (NP_003165.2), African green monkey (ABA56494.1), cow (NP_776615.1), ferret (XP_012904085.1), rabbit (XP_008266240.1), mouse (NP_001334378.1), rat (NP_001101886.2).