Changes in dynamical behavior of the retinoid X receptor DNA-binding domain upon binding to a 14 base-pair DNA half site

The retinoid X receptor (RXR) is a prominent member of the nuclear receptor family of ligand-inducible transcription factors. Many proteins of this fa mily exert their function as heterodimers with RXR as a common upstream par tner. Studies of the DNA-binding domains of several nuclear receptors revea l differences in structure and dynamics, both between the different protein s and between the free- and DNA-bound receptor DBDs. We investigated the di fferences in dynamics between RXR free in solution and in complex with a 14 base-pair oligonucleotide, using H-1 and N-15 relaxation studies. Nano- to picosecond dynamics were probed on N-15, employing Lipari-Szabo analysis w ith an axially symmetric tumbling model to estimate the exchange contributi ons to the transverse relaxation rates. Furthermore, milli- to microsecond dynamics were estimated qualitatively for H-1 and N-15, using CPMG-HSQC and CPMG-T-2 measurements with differential pulse spacing. RXR shows hardly an y nano- to picosecond time-scale internal motion. Upon DNA binding, the ord er parameters show a tiny increase. Dynamics in the milli- to microsecond t ime scale is more prevalent. It is localized in the first and second zinc f ingers of the free RXR. Upon DNA-binding, exchange associated with specific /aspecific DNA-binding of RXR is observed throughout the sequence, whereas conformational flexibility of the D-box and the second zinc finger of RXR i s greatly reduced. Since this DNA-binding induced folding transition occurs remote from the DNA in a region which is involved in protein-protein inter actions, it may very well be related to the cooperativity of dimeric DNA bi nding.