PDBsum entry 4fnc

Hydrolase/DNA

PDB id: 4fnc

Contents

Protein chain

196 a.a.

DNA/RNA

Ligands

HMU

Waters ×38

PDB id:

4fnc

Name:

Hydrolase/DNA

Title:

Human tdg in a post-reactive complex with 5-hydroxymethyluracil (5hmu)

Structure:

G/t mismatch-specific thymine DNA glycosylase. Chain: a. Fragment: human tdg glycosylase domain, unp residues 111-308. Synonym: thymine-DNA glycosylase. Engineered: yes. DNA (28-mer). Chain: c. Engineered: yes. DNA (29-mer).

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: tdg. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Other_details: DNA oligonucleotide synthesis. Other_details: DNA oligonucleotide synthesis

Resolution:

2.49Å R-factor: 0.226 R-free: 0.267

Authors:

H.Hashimoto,S.Hong,A.S.Bhagwat,X.Zhang,X.Cheng

Key ref:

H.Hashimoto et al. (2012). Excision of 5-hydroxymethyluracil and 5-carboxylcytosine by the thymine DNA glycosylase domain: its structural basis and implications for active DNA demethylation. Nucleic Acids Res, 40, 10203-10214. PubMed id: 22962365

Date:

19-Jun-12 Release date: 19-Sep-12

Protein chain

Q13569  (TDG_HUMAN) -  G/T mismatch-specific thymine DNA glycosylase from Homo sapiens

Seq: 410 a.a.

Struc: 196 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains

C-A-G-C-T-C-T-G-T-A-C-G-T-G-A-G-C-A-G-T-G-G-A-C-A-G-C-T 28 bases

A-G-C-T-G-T-C-C-A-C-T-G-C-T-C-A-AAB-G-T-A-C-A-G-A-G-C-T-G-T 29 bases

Enzyme reactions

• Enzyme class: E.C.3.2.2.29 - thymine-DNA glycosylase.

Nucleic Acids Res 40:10203-10214 (2012)

PubMed id: 22962365

Excision of 5-hydroxymethyluracil and 5-carboxylcytosine by the thymine DNA glycosylase domain: its structural basis and implications for active DNA demethylation.

H.Hashimoto, S.Hong, A.S.Bhagwat, X.Zhang, X.Cheng.

ABSTRACT

The mammalian thymine DNA glycosylase (TDG) is implicated in active DNA demethylation via the base excision repair pathway. TDG excises the mismatched base from G:X mismatches, where X is uracil, thymine or 5-hydroxymethyluracil (5hmU). These are, respectively, the deamination products of cytosine, 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC). In addition, TDG excises the Tet protein products 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) but not 5hmC and 5mC, when paired with a guanine. Here we present a post-reactive complex structure of the human TDG domain with a 28-base pair DNA containing a G:5hmU mismatch. TDG flips the target nucleotide from the double-stranded DNA, cleaves the N-glycosidic bond and leaves the C1' hydrolyzed abasic sugar in the flipped state. The cleaved 5hmU base remains in a binding pocket of the enzyme. TDG allows hydrogen-bonding interactions to both T/U-based (5hmU) and C-based (5caC) modifications, thus enabling its activity on a wider range of substrates. We further show that the TDG catalytic domain has higher activity for 5caC at a lower pH (5.5) as compared to the activities at higher pH (7.5 and 8.0) and that the structurally related Escherichia coli mismatch uracil glycosylase can excise 5caC as well. We discuss several possible mechanisms, including the amino-imino tautomerization of the substrate base that may explain how TDG discriminates against 5hmC and 5mC.