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. 2013 Aug 8;8(8):e71228. doi: 10.1371/journal.pone.0071228

Figure 5. The observed effect of DO on peptide presentation is DO specific and can manifest in complex with DM.

Figure 5

(A) Association of HA(Y308A) peptide to DR1 molecules in the presence of coinfected DM/DO complex (Ni-NTA purified). The peptide binding experiment was performed with no accessory molecules (black squares), with DM (red dots), DM/DO (green triangles), or both DM/DO and DM (blue triangles) over the course of 10 hours. The fluorescence signals (Arbitrary Fluorescence Units) associated with the control samples incubated >10 hours in the absence of DR1 were measured: HA(Y308A) peptide alone, 1140; HA(Y308A)+DM, 894; HA(Y308A)+DM/DO, 1404; HA(Y308A)+DM+DM/DO, 1944. (B) DO was depleted from a DO stock by immunoprecipitation via Ni-NTA followed by Mags.DO5 resin. The depleted sample was used instead of DO in reactions measuring HA(anchorless) peptide/DR complex formation in the presence or absence of DM after 5 hours of incubation. The fluorescence intensity of peptide/DR1 complexes formed in the DO depleted reaction was compared to a reaction containing no DO (left bar in each set of three), and a reaction that contained DO that did not undergo depletion (right bar in each set of three). The experiment is representative of three separate trials. (C) A DO-depleted sample was used instead of DO in a reaction measuring of HA(306–318) peptide/DR complex formation in the presence or absence of DM after 5 hours of incubation. The fluorescence intensity of peptide/DR1 complexes formed in the DO depleted reaction was compared to a reaction containing no DO (left bar in each set of three), and a reaction that contained DO that did not undergo depletion (right bar in each set of three). The experiment is representative of three separate trials.