2] The use of nuclear magnetic resonance in the study of transfer RNA structure in solution

Abstract

The chapter discuses the use of nuclear magnetic resonance (NMR) in the study of transfer RNA structure in solution. At both extremes of the proton NMR range there are a manageable number of usually resolved resonances, and it is from these regions of the spectrum that the most detailed, interpretable information on tRNA structure in solution is obtained. The high-field region (between 0 and -4 ppm from DSS) contains aliphatic protons from modified bases, for example, the methyl resonances of methylated nucleotides or the methylene resonances of dihydrouridine. These are nonexchangeable; therefore, they can be studied in either H 2 O or DzO solutions. The low-field region (between -11 and -15 ppm) contains the imino ring NH protons, which are involved in the hydrogen bonding of complementary base pairs. This region of the spectrum is extremely informative in that each base pair contains only one ring NH hydrogen bond; therefore the number of low- field resonances directly reveals the number of stable base pairs in solution. Low-field proton spectroscopy cannot be carried out in D 2 O solutions because of the solvent exchange of the hydrogen-bonded ring NH protons. For the same reason, the rapid exchange of unpaired ring NH protons in contact with water effectively eliminates these protons from the low-field spectrum by time averaging into the H 2 O peak at ca. -4.7ppm. A distinct disadvantage of this form of NMR spectroscopy is the presence of the large (110 M) H 2 O proton peak; however, this is amply compensated by the ability to monitor a single natural reporter group from each base pair in a tRNA molecule.