AlkA protein is the third Escherichia coli DNA repair protein excising a ring fragmentation product of thymine.
Various forms of oxidative stress lead to the formation of damaged bases including N-(2-deoxy-beta-D-erythro-pentofuranosyl)-N-3-(2R-hydroxyisobutyric acid)-urea or alphaRT, the fragmentation product of thymine formed from 5R-thymidine C5-hydrate upon hydrolysis. It was shown that alphaRT is excised by Escherichia coli Fpg and Nth proteins. Here we report that when present in DNA, alphaRT is, in addition, a substrate for the E. coli AlkA protein with an apparent K(m) value of congruent with170 nM. alphaRT positioned opposite T, dG, dC, and dA were efficiently excised by AlkA protein from duplex oligodeoxynucleotides in the following order: dA approximately T >> dC approximately dG. This is the first example of the excision of a ring opened form of a pyrimidine by AlkA protein and also the first example where the same DNA base lesion is excised by three different DNA glycosylases of the base excision repair pathway. The present results suggest possible structural similarity of the active site between E. coli AlkA, Fpg, and Nth proteins.
Privezentzev
C V
CV
Groupe Reparation des lesions Radio- et Chimio-Induites, UMR 8532 CNRS, Institut Gustave Roussy, 94805 Villejuif Cedex, France.
Saparbaev
M
M
Sambandam
A
A
Greenberg
M M
MM
Laval
J
J
D020121:5' Untranslated Regions / Q000378:metabolism; D000818:Animals; D020029:Base Pairing; D053938:DNA Fragmentation; D045647:DNA Glycosylases; D004260:DNA Repair; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009153:Mutagens / Q000378:metabolism; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000378:metabolism*; D009692:Nucleic Acid Heteroduplexes / Q000378:metabolism; D009841:Oligonucleotides / Q000378:metabolism; D051381:Rats; D013379:Substrate Specificity; D013936:Thymidine / Q000031:analogs & derivatives* / Q000378:metabolism; D013941:Thymine / Q000378:metabolism*; D014508:Urea / Q000031:analogs & derivatives* / Q000378:metabolism
D020121:5' Untranslated Regions; D009153:Mutagens; C110651:N-(2-deoxy-erythro-pentofuranosyl)urea; D009692:Nucleic Acid Heteroduplexes; D009841:Oligonucleotides; C097218:thymidine C5-hydrate; D014508:Urea; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C076131:DNA-3-methyladenine glycosidase II; D013941:Thymine; D013936:Thymidine