Thermodynamics and kinetics for base-pair opening in the P1 duplex of the Tetrahymena group I ribozyme

Abstract

The thermodynamics and kinetics for base-pair opening of the P1 duplex of the Tetrahymena group I ribozyme were studied by NMR hydrogen exchange experiments. The apparent equilibrium constants for base pair opening were measured for most of the imino protons in the P1 duplex using the base catalysts NH3, HPO42− or TRIS. These equilibrium constants were also measured for several modified P1 duplexes, and the C-2·G23 base pair was the most stable base pair in all the duplexes. The conserved U-1·G22 base pair is required for activity of the ribozyme and the data here show that this wobble base pair destabilizes neighboring base pairs on only one side of the wobble. A 2′-OMe modification on the U-3 residue stabilized its own base pair but had little effect on the neighboring base pairs. Three base pairs, U-1·G22, C-2·G23 and A2·U21 showed unusual equilibrium constants for opening and possible implications of the opening thermodynamics of these base pairs on the undocking rates of the P1 helix with catalytic core are discussed.

The Zβ domain of human DAI binds to Z-DNA via a novel B-Z transition pathway

“…Hydrogen exchange rate constants of the imino protons of free CG6 and the CG6-hZb DAI complexes at various P/N ratios were determined by a water magnetization transfer experiment [17][18][19][20][21] …”

“…where I 0 and I(t) are the peak intensities of the imino proton in the water magnetization transfer experiments at times zero and t, respectively, and R 1a and R 1w are the apparent longitudinal relaxation rate constants for the imino proton and water, respectively; these parameters were measured with semi-selective inversion recovery 1D NMR experiments [17,18].…”

Sequence discrimination of the Zα domain of human ADAR1 during B–Z transition of DNA duplexes

“…The apparent relaxation rate constant of water (R 1w ) was determined by a selective inversion recovery experiment, using a DANTE sequence for selective water inversion [24]. R 1a and R 1w were determined by curve fitting of the inversion recovery data to the appropriate single-exponential function.…”

“…where I 0 and I(t) are the peak intensities of the imino proton in the water magnetization transfer experiments at times zero and t, respectively, and R 1a and R 1w are the apparent longitudinal relaxation rate constants for the imino proton and water, respectively, measured in semi-selective inversion recovery 1D NMR experiments [24,25].…”

“…4) [24][25][26][27][28]. The protein binding to DNA duplex could change the rate constant and/or the equilibrium constant for base-pair opening (see Supplementary material).…”