Skip to main content
. 2017 Jan 26;85(2):e00788-16. doi: 10.1128/IAI.00788-16

FIG 5.

FIG 5

Secretion of HpaH is independent of the T2S systems and OMVs. (A) HpaH is secreted in the absence of functional T2S systems. Strains 85-10 (wt), 85-10hpaHoof (hpaHoof), and 85-10ΔEEE (ΔT2S), which is deficient in T2S, containing vector pBRM (EV) or encoding HpaH–c-Myc (HpaH) were incubated in NYG medium. Total cell extracts (TE) and culture supernatants (SN) were analyzed by immunoblotting using a c-Myc epitope-specific antibody. (B) The secreted cleavage product of HpaH is degraded by proteinase K. Strain 85* encoding HpaH–c-Myc was incubated in secretion medium. TE and SN were incubated in the presence (+) or absence (−) of proteinase K as indicated and analyzed by immunoblotting using antibodies specific for the c-Myc epitope and the periplasmic HrpB1 protein. As a control, the experiment was performed with strain 85-10 encoding the T2S substrate XCV4360–c-Myc, which was previously localized in OMVs. For the analysis of secreted XCV4360–c-Myc, bacteria were grown in NYG medium.