Kinetic and thermodynamic analysis of 9-cis-retinoic acid binding to retinoid X receptor alpha

The interaction of retinoid X receptor alpha with 9-cis-retinoic acid was s tudied using stopped-flow fluorescence spectroscopy. Transient kinetic anal yses of this interaction suggest a two-step binding mechanism involving a r apid, enthalpically driven pre-equilibrium followed by a slower, entropical ly driven reaction that may arise from a conformational change within the l igand binding domain of the receptor. The assignment of this kinetic mechan ism was supported by agreement between the overall equilibrium constant, K- ov, derived from kinetic studies with that determined by equilibrium fluore scence titrations. Although these analyses do not preclude ligand-induced a lteration in the oligomerization state of the receptor in solution, the sim plest model that can be applied to these data involves the stoichiometric i nteraction of 9-cis-retinoic acid with retinoid X receptor alpha monomers.