FIG. 4.

Essential residues required for the formation of pilin-sortase intermediates. Cells of the wild-type (WT) strain and its isogenic derivative carrying deletions of spaA and srtA or the derivative expressing various SrtA mutants grown in HIB were fractionated into culture medium (M), cell wall (W), and protoplast fractions (P) as described in the legend to Fig. 2. Proteins were separated on 4 to 12% Tris-glycine gradient gels and detected by immunoblotting them with the specific antisera α-SrtA (A) and α-SpaA (B). The positions of molecular mass markers (kDa) and high-molecular-weight polymers (_HMW) are indicated.