The apparent on and off rate constants for binding of theophylline to its RNA aptamer in the absence of Mg2+ were determined here by 2D H-1-H-1 ZZ-exchange NMR spectroscopy. Analysis of the buildup rate of the exchange cross peaks for several base-paired imino protons in the RNA yielded an apparent k(on) of 600 M-1 s(-1). This small apparent k(on) results because the free RNA exist as a dynamic equilibrium of inactive states rapidly interconverting with a tow population of active species. The data found here indicate that the RNA aptamer employs a conformational selection mechanism for binding theophylline in the absence of Mg2+. The kinetic data found here also explain a very unusual property of this RNA-theophylline system: slow exchange on the NMR chemical shift time scale for a weakly binding complex. To our knowledge, it is unprecedented to have such a weakly binding complex (K-d approximate to 3.0 mM at 15 degrees C) show slow exchange on the NMR chemical shift time scale, but the results clearly demonstrate that stow exchange and weak binding are readily rationalized by a small k(on). Comparisons with other ligand-receptor interactions are presented.