CFP10 discriminates between nonacetylated and acetylated ESAT‐6 of Mycobacterium tuberculosis by differential interaction

ESAT‐6 (the 6 kDa early secreted antigenic target) protein species in short‐term culture filtrate of Mycobacterium tuberculosis were separated in a 4–5 narrow range p I gradient two‐dimensional gel electrophoresis (2‐DE). Eight ESAT‐6 protein species were analyzed in detail by peptide mass fingerprinting matrix‐assisted laser desorption/ionization‐mass spectrometry as well as by electrospray ionization‐tandem mass spectrometry. An N ‐terminal Thr acetylation was identified in four species and a C ‐terminal truncation was identified in two species. In 2‐DE blot overlay assays, the recombinant 10 kDa culture filtrate protein (CFP10) discriminated N ‐terminal acetylated and nonacetylated ESAT‐6 by differential interaction, whereas removal of the C ‐terminal 11 residues of ESAT‐6 had no effects thereon. This example shows that the access to the protein species level can be a prerequisite to understand regulation of protein‐protein interaction.