title: Recognition and processing of a new repertoire of DNA substrates by human 3-methyladenine DNA glycosylase (AAG).
abstract: The human 3-methyladenine DNA glycosylase (AAG) recognizes and excises a broad range of purines damaged by alkylation and oxidative damage, including 3-methyladenine, 7-methylguanine, hypoxanthine (Hx), and 1,N(6)-ethenoadenine (epsilonA). The crystal structures of AAG bound to epsilonA have provided insights into the structural basis for substrate recognition, base excision, and exclusion of normal purines and pyrimidines from its substrate recognition pocket. In this study, we explore the substrate specificity of full-length and truncated Delta80AAG on a library of oligonucleotides containing structurally diverse base modifications. Substrate binding and base excision kinetics of AAG with 13 damaged oligonucleotides were examined. We found that AAG bound to a wide variety of purine and pyrimidine lesions but excised only a few of them. Single-turnover excision kinetics showed that in addition to the well-known epsilonA and Hx substrates, 1-methylguanine (m1G) was also excised efficiently by AAG. Thus, along with epsilonA and ethanoadenine (EA), m1G is another substrate that is shared between AAG and the direct repair protein AlkB. In addition, we found that both the full-length and truncated AAG excised 1,N(2)-ethenoguanine (1,N(2)-epsilonG), albeit weakly, from duplex DNA. Uracil was excised from both single- and double-stranded DNA, but only by full-length AAG, indicating that the N-terminus of AAG may influence glycosylase activity for some substrates. Although AAG has been primarily shown to act on double-stranded DNA, AAG excised both epsilonA and Hx from single-stranded DNA, suggesting the possible significance of repair of these frequent lesions in single-stranded DNA transiently generated during replication and transcription.
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
authors.forename: Chun-Yue I
authors.initials: CY
authors.lastname: Lee
authors.forename: James C
authors.initials: JC
authors.lastname: Delaney
authors.forename: Maria
authors.initials: M
authors.lastname: Kartalou
authors.forename: Gondichatnahalli M
authors.initials: GM
authors.lastname: Lingaraju
authors.forename: Ayelet
authors.initials: A
authors.lastname: Maor-Shoshani
authors.forename: John M
authors.initials: JM
authors.lastname: Essigmann
authors.forename: Leona D
authors.initials: LD
authors.lastname: Samson
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D002384:Catalysis; D020134:Catalytic Domain; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage; D045647:DNA Glycosylases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D006147:Guanine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D009841:Oligonucleotides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017384:Sequence Deletion; D013379:Substrate Specificity
chemical_list: D004277:DNA, Single-Stranded; D009841:Oligonucleotides; C004981:1,N(6)-ethenoadenine; C017515:1-methylguanine; C085534:1,(N2)-ethenoguanine; D006147:Guanine; C008450:7-methylguanine; D004247:DNA; C021565:3-methyladenine-DNA glycosylase; D045647:DNA Glycosylases; D000225:Adenine