This proposal addressed several fundamental problems in the chemistry and biochemistry of biologically-important furanose and furanose-containing compounds. The experimental approach involves the combination of stable isotopic enrichment, modern multi-pulse 1D and 2D NMR spectroscopy, and computational chemistry (ab initio molecular orbital calculations). New/improved methods (chemical and enzymic) will be developed to introduce stable isotopes (13C,2H) into carbohydrates, with particular focus on furanose sugars. By preparing systematically a variety of furanose structures, we aim to: A) Study the tautomeric composition of these (13C)-enriched furanose rings in aqueous solution as a function of ring structure/configuration. B) Study the conformational/dynamic properties of these (13C)- enriched furanoses as a function of ring structure and configuration. This will be accomplished by interpreting long- range 1H-1H, 13C-1H and 13C-13C coupling constants obtained from high-resolution NMR spectra. Theoretical input to these studies will be supplied by molecular orbital calculations (geometry optimization). C) Study how furanose ring structure/configuration, and other factors, affect the rates of ring-opening and ring-closing reactions in solution (furanose anomerization). For example, the effect of ring alkylation on these rates (Thorpe-Ingold effects) will be examined. D) Study how furanose ring conformation/dynamics are altered when incorporated into oligonucleotides; specific-sequence oligonucleotides containing stable isotopes will be used in this work. The proposed studies are a prerequisite to the pursuit of the longer-term objectives of determining the molecular basis for the binding specificity between nucleic acids and proteins, and on elucidating the effect of sugar tautomerization on metabolic regulation.
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