Proton FT NMR Studies of tRNA Structure and Dynamics

Abstract

For several years we have been studying tRNA with the primary goal of learning something about its points of flexibility. We have developed a method for measuring the out-exchange rate, over a wide temperature range, of each exchangeable proton whose nuclear magnetic resonance (NMR) line can be resolved or partially resolved. Our hope is that this will reflect the rate at which portions of the structure open, or open partially, to expose these protons to the solvent and that this information will be useful in unraveling modes of tRNA interactions with proteins and other nucleic acids.

More recently we have developed the double-resonance nuclear Overhauser effect (NOE) method for exchangeable resonances in tRNA. If there is a resonance at a frequency f₂ in the spectrum that is at least partially resolved, then, as we will explain, this method allows us to pick out resonances at f₃, f₄, etc., of those one, two, or three protons that are closest to the semiresolved proton (usually within 3.5 Å) that resonates at f₂. This multiplies the information content concerning the original resonance f₂, helping in its identification. When identification is possible for the resonance at f₂, then, generally, the linked resonances at f₃ and f₄ can be assigned and used to extract further conformation and dynamic information. The resonances at f₃ and f₄ are usually in the confused central region of the spectrum and would otherwise be completely useless and unobservable.

We explain the technology behind these experiments and then show how the...