Reinvestigation of the molecular influence of hypoxanthine on the DNA cleavage efficiency of restriction endonucleases BglII, EcoRI and BamHI.
Hypoxanthine (Hyp), a deaminated base of adenine (Ade), can be employed as a good probe molecule to reveal the significance of the minor groove of guanine (Gua) in biomolecular interactions because Hyp possesses a similar structure to Gua lacking its 2-amino group. In this study, we examined cleavage efficiencies of restriction endonuclease enzymes on DNA substrates with Hyp in their recognition sequences. As a substrate for BglII, EcoRI and BamHI, 24-mer DNA oligomer with Hyp (in place of Gua) was prepared together with its complementary sequences with cytosine (Cyt) or thymine (Thy) as the counter base. At 37 degrees C incubation for 1 h, BglII and EcoRI showed higher DNA cleavage reactivity on Hyp-containing DNA substrates than on normal ones, whereas BamHI showed lower values on Hyp-containing substrates. Such high cleavage performance of BglII and EcoRI on Hyp-containing DNA substrates is in contrast to the results obtained 20 years ago, in which short DNA substrates (8- or 10-mer) and low reaction temperatures (15-20 degrees C) were employed. These new results suggest that the lack of the exocyclic 2-amino group of Gua could contribute to enhanced recognition access of BglII and EcoRI to DNA substrates.
Institute of Advanced Energy, Kyoto University, Gokasho, Uji, Japan.
Akihiro
A
Doi
Seung Pil
SP
Pack
Tsutomu
T
Kodaki
Keisuke
K
Makino
D001426:Bacterial Proteins / Q000737:chemistry*; D001483:Base Sequence; D053837:DNA Cleavage; D004262:DNA Restriction Enzymes / Q000737:chemistry / Q000378:metabolism*; D015245:Deoxyribonuclease BamHI / Q000737:chemistry / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000737:chemistry / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006860:Hydrogen Bonding; D019271:Hypoxanthine / Q000737:chemistry*; D007700:Kinetics; D008969:Molecular Sequence Data; D015394:Molecular Structure
D001426:Bacterial Proteins; D019271:Hypoxanthine; D004262:DNA Restriction Enzymes; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI; C024404:BglII endonuclease; D015252:Deoxyribonucleases, Type II Site-Specific