Figure 5. The last six amino acids of EsxD are important for secretion of EsxA and EsxC.

Bacterial cultures of the S. aureus esxD mutant harboring either the empty vector or vector with wild type esxD (pesxD_WT), truncated esxD (pesxD_1-99) or the esxD variant with double alanine substitutions (pesxD_{Y100A, E104A}) were grown to OD_600nm of 1.0 and separated into cell and medium fractions. Proteins were precipitated with trichloroacetic acid, separated by SDS-PAGE and detected by immunoblotting with specific antibodies (α-EsxA, α-EsxC, α-EsxB, α-EsxD, α-EssB, α-EssD, and α-Hla or α-L6 for fractionation and loading controls).