FIG 5.
HNP-1, but not full length preproHNP-1, interacts with RecA. (A) Global distribution of HNP-1-interacting proteins in biological replicate samples. The proteins with log10 intensity ratio >4 were selected for further analysis. HNP-1 itself and heavily enriched RecA are labeled with red arrows. (B) The His6 tagged HNP-1 interacted with RecA. HNP-1 was enriched through a 6x His-tag for purification with Ni-NTA beads and probed with RecA antibody. (C) HNP-1, but not full length 6x His-tagged preproHNP-1, interacted with RecA. Cells expressing HNP-1 were collected at different time points as indicated. RecA was immunoprecipitated with an anti-RecA antibody, followed by Western blotting using an anti-HNP-1 antibody. (D) HNP-1 inhibition of ssDNA-stimulated RecA ATPase activity. ATPase activity was measured by monitoring the release of inorganic phosphate using a malachite green phosphate detection assay. (E) HNP-1 inhibited RecA-ssDNA binding. RecA-bound ssDNA and free ssDNA were resolved using an agarose gel. (F) Quantification of RecA mRNA in MRSA by real-time RT-PCR. (G) Quantification of RecA in MRPA mRNA by real-time RT-PCR. Data were analyzed using a two-tailed Student's t test and are plotted as the mean ± SD for each condition. *, P < 0.05 and ***, P < 0.001.