Bovine spleen, phosphodiesterases from

Fig. 2.8. Nearest neighbour analysis and quantitative depurination analysis of a defined product from a primed synthesis reaction. When radioactive dATP (or dGTP) is used in the primed synthesis, depurination analysis will yield pyrimidine tracts each of which terminate in a radioactive 3 -phosphate. Thus only those depurination products which lie 5 -adjacent to the labelled nucleotide will be labelled. Each depurination product will be labelled to the same specific activity thus greatly simplifying the quantitation. Digestion of the labelled product with a mixture of micrococcal nuclease and bovine spleen phosphodiesterase yields the nucleoside 3 -monophosphates. Identification of the labelled products (by paper electrophoresis at pH 3.S) gives the nearest neighbours to the labelled substrate.

E. Phosphodiesterases from Snake Venom and Bovine Spleen. 133... 

We have also studied the stereochemical outcomes of the reactions catalyzed by the phosphodiesterase from snake venom (99) and from bovine spleen (101). The former enzyme catalyzes the hydrolysis of esters of 5 -nucleotides (free 3 -hydroxyl group), and the latter enzyme catalyzes the hydrolysis of esters of 3 -nucleotides (free 5 -hydroxyl group). No structural information of mechanistic consequence is available for these enzymes. 

The incorporation of the 5R and 5S diastereoisomers of the 5-hydroxyhydan-toin (derived from oxidative damage of dC) derivative (89) has been achieved using phosphoramidite chemistry and mild deprotection. Whilst the ODNs containing either diastereoisomer were digested with nuclease Pi, the hydantoin derivatives were resistant to digestion by calf spleen and bovine intestinal mucosa phosphodiesterases. 

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