Recognition by restriction endonuclease EcoRI of deoxyoctanucleotides containing modified sugar moieties

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Abstract
The deoxyribooctanucleotide d(G‐G‐A‐A‐T‐T‐C‐C), containing the recognition sequence for EcoRI, d(G‐A‐A‐T‐T‐C), and analogs containing modified sugar moieties were tested for their activity in cleavage with EcoRI. These analogs, with replacement in the third position from the 5′ end, were synthesized using 9‐β‐d‐arabinosyladenine (aA), 2′‐deoxy‐2′‐fluoroadenosine (Afl) and adenosine (rA). Duplex formation by the three analogs was confirmed by measurements of ultraviolet/temperature profiles. It was found that EcoRI cleaved these duplexes less efficiently than d(G‐G‐A‐A‐T‐T‐C‐C). The adenosine‐containing analog d(G‐G)‐rA‐d(A‐T‐T‐C‐C) was cleaved much more slowly than d(G‐G)‐aA‐d(A‐T‐T‐C‐C) and d(G‐G‐Afl‐A‐T‐T‐C‐C). The corresponding ribooctamer G‐G‐A‐A‐U‐U‐C‐C showed a higher melting temperature than the deoxyoctamers but its duplex was not cleaved by this enzyme. An analog with 2′‐deoxy‐2′‐fluoroguanosine at the second position was cleaved by the endonuclease faster than the natural deoxyoctamer.

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