Fig 5.

Ply truncation mutants reveal that domain 2 of Ply may be required for export. (A) Model of Ply three-dimensional structure. The model was made with ModWeb software and manipulated with MacPyMol software. Domains are color coded: blue, domain 1; green, domain 2; red, domain 3; yellow, domain 4. (B) Schematic of primary sequences of Ply and truncation mutants. Domains are color coded as in panel A. Truncations of Ply were made from the N and C termini, tagged with an HA epitope at the C terminus of each construct, and crossed into the ply locus on the S. pneumoniae chromosome. Strains were grown to mid-exponential phase, fractionated into cell wall and protoplast compartments, and assayed for the presence of Ply and CodY by Western blotting. (C) N-terminal truncation mutants. The band in the supernatant lane for the N-4 truncation (shown by the asterisk) is of incorrect size and is unexplained. (D) C-terminal truncation mutants. Equal cell equivalents were loaded on the gel. Sup, culture supernatant; CW, cell wall; Prt, protoplast.