A DNA enzyme with N-glycosylase activity.
In vitro evolution was used to develop a DNA enzyme that catalyzes the site-specific depurination of DNA with a catalytic rate enhancement of about 10(6)-fold. The reaction involves hydrolysis of the N-glycosidic bond of a particular deoxyguanosine residue, leading to DNA strand scission at the apurinic site. The DNA enzyme contains 93 nucleotides and is structurally complex. It has an absolute requirement for a divalent metal cation and exhibits optimal activity at about pH 5. The mechanism of the reaction was confirmed by analysis of the cleavage products by using HPLC and mass spectrometry. The isolation and characterization of an N-glycosylase DNA enzyme demonstrates that single-stranded DNA, like RNA and proteins, can form a complex tertiary structure and catalyze a difficult biochemical transformation. This DNA enzyme provides a new approach for the site-specific cleavage of DNA molecules.
Sheppard
T L
TL
Departments of Chemistry and Molecular Biology, The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Ordoukhanian
P
P
Joyce
G F
GF
D002384:Catalysis; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D045647:DNA Glycosylases; D004260:DNA Repair; D003849:Deoxyguanosine / Q000378:metabolism; D019020:Directed Molecular Evolution; D008956:Models, Chemical; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D012641:Selection, Genetic
D009838:Oligodeoxyribonucleotides; D004247:DNA; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; D003849:Deoxyguanosine