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. 2012 Dec 3;7(12):e50490. doi: 10.1371/journal.pone.0050490

Figure 2. Binding affinities of the interactions between p97/VCP and its adaptor proteins.

Figure 2

p97/VCP was immobilized on a CM5 sensorchip surface, using amine coupling procedure, and a concentration series of p47 (two fold dilutions from 1.92 μM) or UN (two fold dilutions from 1.88 μM) were injected over the immobilized surface at a flow rate of 30 μl/min, at 25°C, using PBS as the sample and running buffer. Sensorgrams and the corresponding interaction maps for the p97/VCP-UN interaction without ATP (A), with ATP during UN association and dissociation (B) and for p97/VCP-p47 interactions when p47 is immobilized (C) and when p97/VCP is immobilized (D) are shown. Interaction Map Analysis shows that the interaction between UN and p97/VCP is complex, with a major component with ∼5 μM affinity, and a second component with ∼400 nM affinity. When ATP is added, the affinity of the second component increases to 100 nM. For the interaction between p97/VCP and p47, only one kinetic component was found, and the interaction could be fitted to a Langmuir 1∶1 interaction model, with ∼31.3 nM affinity when p47 is immobilized and p97 injected across (C) or ∼5 μM affinity when p97 is immobilized and p47 injected across (D).