Probes for Studies of Nucleotide-Binding Proteins

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DESCRIPTION (provided by applicant): This project focuses on the development of a novel class of transglycosidically tethered 5'-nucleotides by synthesis and physicochemical evaluation. A single general synthetic strategy for the tethering of any 5'-nucleotide of interest has already been developed. It was used successfully to prepare the first two tethered 5'-nucleotide targets. The specific nature of this new tether and its mode of attachment to a nucleotide framework allow an unprecedented degree of 5'-nucleotide biomimicry according to the conformational disposition of the protein-bound ligands in the Protein Data Bank. The tethered 5'-nucleotides are for all practical purposes chemically identical to their natural counterparts but are restricted to a limited set of spatial conformations. Further development will confirm their highly biomimetic features and demonstrate their utility to biomedical investigations. Synthesis of a variety of carefully selected targets, and extensive characterization by NMR spectroscopy and X-ray crystallography are the specific activities to be undertaken in this work. Conformational restriction can generate valuable probes of protein-ligand structure-function relationships. Just two bond rotations determine most of the molecular topography of 5'-nucleotides: the C4'-C5' bond and the glycosidic one (C1'-N1 for pyrimidines and C1'-N9 for purines), and these are restricted simultaneously by the new tether. The desirable features of a transglycosidic nucleotide-tethering motif include restriction of the glycosidic bond in an anti conformation, preservation of the biorecognition-critical hydrogen bonding sites of the heterocyclic aglycon, retention of the functionalizable carbohydrate hydroxyl groups, and a design that can be used on both the pyrimidine and the purine nucleoside frameworks. The tethering motif featured in this proposal meets all these criteria, and future use of the tethered targets could contribute to an understanding of conformation/binding/function relationships in many biochemically important 5'-nucleotide-binding proteins.