FIG 4.
NPB on PspA and PspC binds to LDH. (A) Immunoblots of total bacterial cell lysates using anti-NPB monoclonal antibody PR-1A4.7 with total bacterial cell lysate was used to detect the NPB domain in WU2, TIGR4, and EF3030. (B) Spn WU2, TIGR4, or EF3030 was incubated with FITC-conjugated LDH. LDH binding to pneumococci was measured using flow cytometry. This experiment was performed two times with identical results. (C) Immunoblots using NPB monoclonal antibody of total Spn bacterial cell lysates from designated clinical isolates. The presence of PspA or PspC containing the NPB as inferred from publicly available genomic sequence data is indicated on the bottom. (D) The ability of these isolates to bind rabbit LDH-A was also measured using flow cytometry (n = 3). (E) EF3030, EF3030 isogenic pspA mutant (EF3030-01), EF3030 expressing PspAWU2 (EF3030-04), and EF3030 expressing PspAWU2 without NPB (EF3030-05) were incubated with FITC-conjugated rabbit LDH-A. The LDH-A binding to pneumococci was measured by flow cytometry. (F) Spn WU2, S. aureus, and urinary tract-pathogenic E. coli (UP E. coli) were incubated with FITC-conjugated rabbit LDH-A. The LDH binding to pneumococci was measured by flow cytometry. This experiment was performed two times with similar results.