The influence of sequences adjacent to the recognition site on the cleavage of oligodeoxynucleotides by the EcoRI endonuclease

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Abstract
The influence of the nucleotide sequence adjacent to the recognition site on the rate of cleavage of DNA by the restriction endonuclease EcoRI was investigated. For this purpose 2 decadeoxynucleotides, d(G-G-G-A-A-T-T-C-T-T) (Ia) and d(A-A-G-A-A-T-T-C-C-C-) (Ib) were synthesized. The duplex Ia .cntdot. Ib is cleaved by EcoRI preferentially in the dA-rich strand (.apprx. 10 times over the dG-rich strand). The individual nucleotides Ia and Ib are also cleaved by EcoRI, Ib at a higher rate than Ia and both at a lower rate than Ia .cntdot. Ib. The temperature dependence of the reaction shows that only double-stranded oligodeoxynucleotides are substrates for the EcoRI endonuclease. Oligomers of d(G-G-A-A-T-T-C-C) were synthesized, which contain 2, 3 and 4 EcoRI sites, respectively. These oligodeoxynucleotides are preferentially cleaved at the sites next to the 5'' end, where the recognition site is only flanked by 1 dG .cntdot. dC base pair, in contrast to the other sites which are flanked by 3 such pairs. Apparently, sequences adjacent to the recognition site influence the rate of cleavage; dA .cntdot. dT base pairs enhance the dG .cntdot. dC base pairs slow down the hydrolytic activity of the EcoRI endonuclease.

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