FLUORODEOXY SUGAR ANALOGS OF 2',5'-OLIGOADENYLATES AS PROBES OF HYDROGEN-BONDING IN ENZYMES OF THE 2-5A SYSTEM

In order to understand further the contribution of the 3’-hydroxyl groups of 2-5A (ppp5’(A2’p(n)A) to its interaction with RNase L, we synthesized a series of analogues in which the 3’-hydroxyl moiety was replaced by fluorine to yield 3’-fluoro-3’-fluoro-3’-deoxyadenosine (A(F)). These included ppp5’A2’p5’A2’p5A(F), ppp5’A2’p5’A(F)2’p5’A, ppp5’A(F)2’p5’A2’p5’A, ppp5’A(F)2’p5’A(F0, ppp5’A2’p5’A(F)2’p5’A(F), ppp5’A(F)2’p5’A2’p5’A(F), and the corresponding monophosphates. When these oligomers were evaluated for their ability to activate RNase L from various sources, we found that the replacement of the second from the 5’-terminus adenosine residue of 2-5A with the fluoro analogue caused major reductions in activity. We conclude that the hydroxyl group of this second or middle nucleotide residue of 2-5A trimer may act as a hydrogen bond donor to an acceptor group in RNase L and that this hydrogen bond may be key to what we presume may be a critical conformational change required for nuclease activity of RNase L. We have also found that this substitution of fluorine for hydroxyl in the second or penultimate residue of 2-5A trimer results in an oligomer with a 2’,5’-phosphodiesterase sensitivity comparable to that of 2-5A itself. When viewed in terms of earlier experiments, these results suggest that the role of the 3’-OH group of the penultimate nucleotide of 2-5A may be to anchor the substrate to the phosphodiesterase through its action as a hydrogen bond receptor. © 1993 by Academic Press, Inc.