Structure and activity of a thermostable thymine-DNA glycosylase: evidence for base twisting to remove mismatched normal DNA bases

• Published in: Journal of molecular biology 2002-01, Vol.315 (3), p.373-384
• Main Authors: Mol, Clifford D, Arvai, Andrew S, Begley, Thomas J, Cunningham, Richard P, Tainer, John A
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Base Pair Mismatch - genetics; base twisting; Binding Sites; Carbon-Oxygen Lyases - chemistry; Carbon-Oxygen Lyases - metabolism; Crystallography, X-Ray; Deoxyribonuclease (Pyrimidine Dimer); DNA - chemistry; DNA - genetics; DNA - metabolism; DNA glycosylase; DNA mismatch; DNA repair; DNA Repair - genetics; DNA-(Apurinic or Apyrimidinic Site) Lyase; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Endodeoxyribonucleases - chemistry; Endodeoxyribonucleases - genetics; Endodeoxyribonucleases - metabolism; Methanobacteriaceae - enzymology; Methanobacteriaceae - genetics; methylation; Models, Molecular; Molecular Sequence Data; Mutation - genetics; Nucleic Acid Conformation; Nucleotides - chemistry; Nucleotides - genetics; Nucleotides - metabolism; Protein Structure, Secondary; Protein Structure, Tertiary; Sequence Alignment; Sequence Homology, Amino Acid; Structure-Activity Relationship; Thermodynamics; Thymine - metabolism; thymine-DNA glycosylase
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1006/jmbi.2001.5264

The repair of T:G mismatches in DNA is key for maintaining bacterial restriction/modification systems and gene silencing in higher eukaryotes. T:G mismatch repair can be initiated by a specific mismatch glycosylase (MIG) that is homologous to the helix-hairpin-helix (HhH) DNA repair enzymes. Here, we present a 2.0 Å resolution crystal structure and complementary mutagenesis results for this thermophilic HhH MIG enzyme. The results suggest that MIG distorts the target thymine nucleotide by twisting the thymine base ∼90° away from its normal anti position within DNA. We propose that functionally significant differences exist in DNA repair enzyme extrahelical nucleotide binding and catalysis that are characteristic of whether the target base is damaged or is a normal base within a mispair. These results explain why pure HhH DNA glycosylases and combined glycosylase/AP lyases cannot be interconverted by simply altering their functional group chemistry, and how broad-specificity DNA glycosylase enzymes may weaken the glycosylic linkage to allow a variety of damaged DNA bases to be excised.