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Nucleic Acids Research, 1986, Vol. 14, No. 8 3463-3474
© 1986

Articles

Role of thymidine residues in DNA recognition by the EcoRI and EcoRV restriction endonucleases

Anja Fliess1, Heiner Wolfes1, Andre Rosenthal2, Konrad Schwellnus3, Helmut Blöcker3, Ronald Frank3 and Alfred Pingoud1

1Abteilung Biophysikalische Chemie Konstanty-Gutschow-Str. 8, D-3000 Hannover 61, FRG 2Akademie der Wissenschaften der DDR, Zentralinstitut fur Molekularbiologie, Abteilung fur Zelldifferenzierung Robert-R6ssle-Str. 10, DDR-1115 Berlin-Buch, GDR 3DNA-Synthese Gruppe, Gesellschaft fur Biotechnologische Forschung mbH Mascheroder Weg 1, D-3300 Braunschweig, FRG

Received February 12, 1986. Accepted March 27, 1986.

We have synthesized a series of oligonucleotides containing the EcoRI (GAATTC) or EcoRV (GATATC) recognition site within which or adjacent to which thymidine was substituted by uridine or derivatives of uridine. The effects of these substitutions on the rate of the EcoRI and EcoRV catalyzed cleavage reaction were investigated. Our results show that most of the substitutions within the site are quite well tolerated by EcoRI.not, however, by EcoRV. We conclude that the thymin residues most likely are not directly involved in the recognition process of the EcoRI reaction. In contrast they are major points of contact.between substrate and enzyme in the EcoRV reaction. The effects of substitutions in the position adjacent to the recognition site is also markedly different for EcoRI and EcoRV. Here, EcoRI seems to be considerably more selective than EcoRV.


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