Fusion protein based epitope mapping of the MPB57 protein from Mycobacterium bovis BCG and its epitope insertion into the native protein

The gene coding for the 12-kDa protein (MPB57) of Mycobacterium bovis BCG has recently been cloned and sequenced (R. Yamaguchi, K. Matsuo, A. Yamazaki, S. Nagai, K. Terasaka, and T. Yamada. 1988. FEBS Lett. 240: 115-117). To map linear B-cell epitopes by beta-galactosidase fusion proteins, we have constructed convenient vectors (pUR278S, pUR288S, and pUR289S) with the SmaI site. Based on recognition by polyclonal antibodies, two epitope regions on the MPB57 protein were identified, both of which corresponded to the amino acid sequences Glu20 to Val45 (26 residues, epitope I region) and Ile78 to Leu86 (9 residues, epitope II). Complementary oligonucleotides encoding epitope II were synthesized, polymerized by a ligase reaction, inserted into the native MPB57 protein gene, and expressed in Escherichia coli, giving rise to epitope-inserted proteins. Their stability and potential uses are described.