Identification and characterization of a human DNA glycosylase for repair of modified bases in oxidatively damaged DNA.
8-oxoguanine (8-oxoG), ring-opened purines (formamidopyrimidines or Fapys), and other oxidized DNA base lesions generated by reactive oxygen species are often mutagenic and toxic, and have been implicated in the etiology of many diseases, including cancer, and in aging. Repair of these lesions in all organisms occurs primarily via the DNA base excision repair pathway, initiated with their excision by DNA glycosylase/AP lyases, which are of two classes. One class utilizes an internal Lys residue as the active site nucleophile, and includes Escherichia coli Nth and both known mammalian DNA glycosylase/AP lyases, namely, OGG1 and NTH1. E. coli MutM and its paralog Nei, which comprise the second class, use N-terminal Pro as the active site. Here, we report the presence of two human orthologs of E. coli mutM nei genes in the human genome database, and characterize one of their products. Based on the substrate preference, we have named it NEH1 (Nei homolog). The 44-kDa, wild-type recombinant NEH1, purified to homogeneity from E. coli, excises Fapys from damaged DNA, and oxidized pyrimidines and 8-oxoG from oligodeoxynucleotides. Inactivation of the enzyme because of either deletion of N-terminal Pro or Histag fusion at the N terminus supports the role of N-terminal Pro as its active site. The tissue-specific levels of NEH1 and OGG1 mRNAs are distinct, and S phase-specific increase in NEH1 at both RNA and protein levels suggests that NEH1 is involved in replication-associated repair of oxidized bases.
Hazra
Tapas K
TK
Sealy Center for Molecular Science and Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX 77555, USA.
Izumi
Tadahide
T
Boldogh
Istvan
I
Imhoff
Barry
B
Kow
Yoke W
YW
Jaruga
Pawel
P
Dizdaroglu
Miral
M
Mitra
Sankar
S
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects; D004249:DNA Damage / Q000235:genetics* / Q000528:radiation effects; D045647:DNA Glycosylases; D004260:DNA Repair; D043382:DNA-Formamidopyrimidine Glycosylase; D030561:Databases, Nucleic Acid; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins; D020869:Gene Expression Profiling; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020161:Physical Chromosome Mapping; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016196:S Phase; D016415:Sequence Alignment; D013379:Substrate Specificity
D029968:Escherichia coli Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D004247:DNA; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C473634:NEIL1 protein, human; D043382:DNA-Formamidopyrimidine Glycosylase; C468610:DNA-formamidopyrimidine glycosylase, E coli