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. Author manuscript; available in PMC: 2009 May 11.
Published in final edited form as: J Mol Biol. 2006 May 19;360(2):421–434. doi: 10.1016/j.jmb.2006.05.014

Table 3.

Comparison of NF-κB(p50/p65) heterodimer and NF-κB(p65/p65) homodimer binding data from SPR and ITC experiments.

Protein Schematic Temperature °C KD-SPR (× 10-9M) KD-ITC (× 10-9M)
p50248-350/p65190-321 graphic file with name nihms-104130-t0009.jpg 37 0.32 ± 0.10 ND
p65190-321/P65190-321 graphic file with name nihms-104130-t0010.jpg 37 0.54 ± 0.08 ND
p50248-350/p651-304 graphic file with name nihms-104130-t0011.jpg 30 ND 125 ± 8
p65248-350/p65190-304 graphic file with name nihms-104130-t0012.jpg 30 ND 320 ± 28
p50248-350/p65190-304 graphic file with name nihms-104130-t0013.jpg 30 ND 330 ± 23
p5039-363/p651-304 graphic file with name nihms-104130-t0014.jpg 30 ND 42 ± 6
p651-304/p651-304 graphic file with name nihms-104130-t0015.jpg 30 ND 51 ± 16

SPR data were fit using the Biaevaluation 4.1 software to a simple Langmuir binding model. Experiments were carried out in triplicate for estimations of errors. In each experiment the amount of immobilized NF-κB was varied in the range of 50 to 400 RU. ITC data were fit to a model for a single set of identical binding sites after the heats of dilution of NF-κB into buffer was subtracted. The stoichiometry for the interaction was found to be consistent with a one-to-one binding model in all cases. For the SPR experiments the experimental temperature depended on the dissociation rate for the interaction. For the very tight binding complexes at the top of the table the dissociation rate was to low to be quantified at temperatures below 37 °C. For ITC experiments the data could not be obtained easily at temperature below 20 °C due to the small ΔHobs and limitations of the instrument.