title: Human Tdp1 cleaves a broad spectrum of substrates, including phosphoamide linkages.
abstract: Human tyrosyl-DNA phosphodiesterase (Tdp1) hydrolyzes the phosphodiester bond between a DNA 3' end and a tyrosyl moiety. In eukaryotic cells, this type of linkage is found in stalled topoisomerase I-DNA covalent complexes, and Tdp1 has been implicated in the repair of such complexes in vivo. We confirm here that the Tdp1 catalytic cycle involves a covalent reaction intermediate in which a histidine residue is connected to a DNA 3'-phosphate through a phosphoamide linkage. Most surprisingly, this linkage can be hydrolyzed by Tdp1, and unlike a topoisomerase I-DNA complex, which requires modification to be an efficient substrate for Tdp1, the native form of Tdp1 can be removed from the DNA. The spinocerebellar ataxia with axonal neuropathy neurodegenerative disease is caused by the H493R mutant form of Tdp1, which shows reduced enzymatic activity and accumulates the Tdp1-DNA covalent intermediate. The ability of wild type Tdp1 to remove the stalled mutant protein from the DNA likely explains the recessive nature of spinocerebellar ataxia with axonal neuropathy. In addition to its activity on phosphotyrosine and phosphohistidine substrates, Tdp1 also possesses a limited DNA and RNA 3'-exonuclease activity in which a single nucleoside is removed from the 3'-hydroxyl end of the substrate. Furthermore, Tdp1 also removes a 3' abasic site and an artificial 3'-biotin adduct from the DNA. In combination with earlier data showing that Tdp1 can use 3'-phosphoglycolate as a substrate, these data suggest that Tdp1 may function to remove a variety of 3' adducts from DNA during DNA repair.
authors.affiliation: Department of Microbiology, School of Medicine, University of Washington, Seattle, Washington 98195-7242, USA.
authors.forename: Heidrun
authors.initials: H
authors.lastname: Interthal
authors.forename: Hong Jing
authors.initials: HJ
authors.lastname: Chen
authors.forename: James J
authors.initials: JJ
authors.lastname: Champoux
mesh_terms: D000483:Alleles; D001369:Axons / Q000473:pathology; D001710:Biotin / Q000737:chemistry; D019921:Biotinylation; D002384:Catalysis; D004247:DNA / Q000737:chemistry; D004260:DNA Repair; D005663:Furans / Q000737:chemistry; D006016:Glycolates / Q000737:chemistry; D006801:Humans; D006868:Hydrolysis; D008956:Models, Chemical; D008957:Models, Genetic; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D019606:Phosphoamino Acids / Q000737:chemistry*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding; D012313:RNA / Q000737:chemistry; D013379:Substrate Specificity; D013997:Time Factors; D014443:Tyrosine / Q000737:chemistry
chemical_list: D005663:Furans; D006016:Glycolates; D010455:Peptides; D019606:Phosphoamino Acids; C018674:tetrahydrofuran; D014443:Tyrosine; D012313:RNA; D001710:Biotin; D004247:DNA; D010727:Phosphoric Diester Hydrolases; C498508:TDP1 protein, human; C013193:phosphoglycolate