FIG. 5.
Strains lacking PrsA2 secrete reduced amounts of LLO and PC-PLC. Bacterial supernatant and surface-associated proteins from the WT or ΔprsA2::erm or ΔprsA2::erm + pPL2-prsA2 mutant were isolated by TCA precipitation (secreted proteins) or boiling in 2% SDS (surface-associated proteins). Sample volumes were adjusted to reflect equivalent bacterial densities (OD600 of ∼1.4). Western analysis of protein fractions subjected to SDS-PAGE was done using rabbit polyclonal anti-LLO, rabbit polyclonal anti-PrsA2, and rabbit polyclonal anti-PC-PLC antibody to detect LLO, PrsA2, and PC-PLC, respectively. Arrows indicate bands of interest (either LLO, PrsA2, or PC-PLC) in each panel. The amount of protein detected for each sample in comparison to the WT lane (set at 1.0) as determined by densitometry is indicated at the bottom of each panel. Lane 1, WT; lane 2, ΔprsA2::erm; lane 3, ΔprsA2::erm + pPL2-prsA2.