Circadian Clock Parameter Measurement: Characterization of Clock Transcription Factors Using Surface Plasmon Resonance

To refine mathematical models of the transcriptional/translational feedback loop in the clockwork of Arabidopsis thaliana, the investigators sought to determine the affinity of the transcription factors LHY, CCA1, and CHE for their cognate DNA target sequences in vitro. Steady-state dissociation constants were observed to lie in the low nanomolar range. Furthermore, the data suggest that the LHY/CCA1 heterodimer binds more tightly than either homodimer and that DNA binding of these complexes is temperature compensated. Finally, it was found that LHY binding to the evening element in vitro is enhanced by both molecular crowding effects and by casein kinase 2-mediated phosphorylation.