Completing the β,γ-CXY-dNTP Stereochemical Probe Toolkit: Synthetic Access to the dCTP Diastereomers and P and F NMR Correlations with Absolute Configurations.

Nucleoside 5'-triphosphate (dNTP) analogues in which the β,γ-oxygen is mimicked by a CXY group (β,γ-CXY-dNTPs) have provided information about DNA polymerase catalysis and fidelity. Definition of CXY stereochemistry is important to elucidate precise binding modes. We previously reported the ()- and ()-β,γ-CHX-dGTP diastereomers (X = F, Cl), prepared via P,C-dimorpholinamide CHCl (, ) and CHF (, ) bisphosphonates (BPs) equipped with an ()-mandelic acid as a chiral auxiliary, with final deprotection using H/Pd. This method also affords the β,γ-CHCl-dTTP (, ), β,γ-CHF (, ), and β,γ-CHCl (, ) dATP diastereomers as documented here, but the reductive deprotection step is not compatible with dCTP or the bromo substituent in β,γ-CHBr-dNTP analogues. To complete assembly of the toolkit, we describe an alternative synthetic strategy featuring ethylbenzylamine or phenylglycine-derived chiral BP synthons incorporating a photolabile protecting group. After acid-catalyzed removal of the ()-(+)-α-ethylbenzylamine auxiliary, coupling with activated dCMP and photochemical deprotection, the individual diastereomers of β,γ-CHBr- (, ), β,γ-CHCl- (, 3), β,γ-CHF-dCTP (, ) were obtained. The β,γ-CH(CH)-dATPs (, ) were obtained using a methyl ()-(-)-phenylglycinate auxiliary. P and F NMR Δδ values are correlated with CXY stereochemistry and p values for 13 CXY-bisphosphonic acids and imidodiphosphonic acid are tabulated.