title: Mechanism of repair of acrolein- and malondialdehyde-derived exocyclic guanine adducts by the α-ketoglutarate/Fe(II) dioxygenase AlkB.
abstract: The structurally related exocyclic guanine adducts α-hydroxypropano-dG (α-OH-PdG), γ-hydroxypropano-dG (γ-OH-PdG), and M1dG are formed when DNA is exposed to the reactive aldehydes acrolein and malondialdehyde (MDA). These lesions are believed to form the basis for the observed cytotoxicity and mutagenicity of acrolein and MDA. In an effort to understand the enzymatic pathways and chemical mechanisms that are involved in the repair of acrolein- and MDA-induced DNA damage, we investigated the ability of the DNA repair enzyme AlkB, an α-ketoglutarate/Fe(II) dependent dioxygenase, to process α-OH-PdG, γ-OH-PdG, and M1dG in both single- and double-stranded DNA contexts. By monitoring the repair reactions using quadrupole time-of-flight (Q-TOF) mass spectrometry, it was established that AlkB can oxidatively dealkylate γ-OH-PdG most efficiently, followed by M1dG and α-OH-PdG. The AlkB repair mechanism involved multiple intermediates and complex, overlapping repair pathways. For example, the three exocyclic guanine adducts were shown to be in equilibrium with open-ring aldehydic forms, which were trapped using (pentafluorobenzyl)hydroxylamine (PFBHA) or NaBH4. AlkB repaired the trapped open-ring form of γ-OH-PdG but not the trapped open-ring of α-OH-PdG. Taken together, this study provides a detailed mechanism by which three-carbon bridge exocyclic guanine adducts can be processed by AlkB and suggests an important role for the AlkB family of dioxygenases in protecting against the deleterious biological consequences of acrolein and MDA.
authors.affiliation: Departments of Biological Engineering, ‡Chemistry, and §Center for Environmental Health Sciences, Massachusetts Institute of Technology , Cambridge, Massachusetts 02139, United States.
authors.forename: Vipender
authors.initials: V
authors.lastname: Singh
authors.forename: Bogdan I
authors.initials: BI
authors.lastname: Fedeles
authors.forename: Deyu
authors.initials: D
authors.lastname: Li
authors.forename: James C
authors.initials: JC
authors.lastname: Delaney
authors.forename: Ivan D
authors.initials: ID
authors.lastname: Kozekov
authors.forename: Albena
authors.initials: A
authors.lastname: Kozekova
authors.forename: Lawrence J
authors.initials: LJ
authors.lastname: Marnett
authors.forename: Carmelo J
authors.initials: CJ
authors.lastname: Rizzo
authors.forename: John M
authors.initials: JM
authors.lastname: Essigmann
mesh_terms: D000171:Acrolein / Q000737:chemistry*; D001894:Borohydrides / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D018736:DNA Adducts / Q000737:chemistry / Q000378:metabolism*; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D003849:Deoxyguanosine / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D008315:Malondialdehyde / Q000737:chemistry*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009841:Oligonucleotides / Q000032:analysis / Q000138:chemical synthesis; D053719:Tandem Mass Spectrometry
chemical_list: D001894:Borohydrides; D018736:DNA Adducts; D004277:DNA, Single-Stranded; D029968:Escherichia coli Proteins; D009841:Oligonucleotides; D008315:Malondialdehyde; D000171:Acrolein; C025364:sodium borohydride; D004247:DNA; D006899:Mixed Function Oxygenases; C466427:AlkB protein, E coli; D003849:Deoxyguanosine