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. 2015 Mar 25;10(3):e0120471. doi: 10.1371/journal.pone.0120471

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© 2015 Solomon et al

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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Fig 5 — A. YopB-YopD complexes secreted in bacterial supernatants of yopD, yopD/pYopD and yopD expressing the different yopD mutants were precipitated by 1 h incubation with YopD mab (clone 248:19) followed by incubation with Dynabeads Protein G (Invitrogen). After 3 washes, beads were resuspended in 2X Laemmli sample buffer, and boiled. The eluted material (IP) and an aliquot of the bacterial supernatants (SN) were resolved in SDS-PAGE. Western blot was performed using anti-YopD and anti-YopB Mabs, and anti-mouse IR680 or IR800 secondary antibodies. Bands corresponding to YopB and YopD are indicated by arrows. Also present in the IP samples are a band originated from the beads (marked with an asterix), and a weak band of unknown origin that migrates right below YopB. B. Signal intensities of immunoprecipitated YopB and YopD were calculated using Odyssey imaging system software (LI-COR Biosience). YopB-YopD interaction for each mutant was calculated as the ratio between immunoprecipitated YopB and YopD. Results were normalized to yopD/pYopD.