PDBsum entry 6n2s

Gene regulation/DNA

PDB id: 6n2s

Contents

Protein chain

326 a.a.

DNA/RNA

Ligands

1GC

Metals

_MG ×2

_NA ×2

Waters ×81

PDB id:

6n2s

Name:

Gene regulation/DNA

Title:

Ternary complex crystal structure of DNA polymerase beta with 5- carboxy-dc (5-cac) at the templating position

Structure:

DNA polymerase beta. Chain: a. Fragment: DNA polymerase beta. Engineered: yes. DNA (5'-d( Cp Cp Gp Ap Cp (1Cc) p Gp Cp Gp Cp Ap Tp Cp Ap Gp C)-3'). Chain: t. Fragment: template strand. Engineered: yes.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: polb. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Organism_taxid: 9606

Resolution:

2.46Å R-factor: 0.211 R-free: 0.290

Authors:

V.K.Batra,S.H.Wilson

Key ref:

M.J.Howard et al. (2019). Molecular basis for the faithful replication of 5-methylcytosine and its oxidized forms by DNA polymerase β. J Biol Chem, 294, 7194-7201. PubMed id: 30885943 DOI: 10.1074/jbc.RA118.006809

Date:

14-Nov-18 Release date: 27-Mar-19

Protein chain

P06746  (DPOLB_HUMAN) -  DNA polymerase beta from Homo sapiens

Seq: 335 a.a.

Struc: 326 a.a.

DNA/RNA chains

C-C-G-A-C-1CC-G-C-G-C-A-T-C-A-G-C 16 bases

G-C-T-G-A-T-G-C-G-C 10 bases

G-T-C-G-G 5 bases

Enzyme reactions

• Enzyme class 1: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 2: E.C.4.2.99.- - ?????
• Enzyme class 3: E.C.4.2.99.18 - DNA-(apurinic or apyrimidinic site) lyase.
• Reaction: 2'-deoxyribonucleotide-(2'-deoxyribose 5'-phosphate)- 2'-deoxyribonucleotide-DNA = a 3'-end 2'-deoxyribonucleotide-(2,3- dehydro-2,3-deoxyribose 5'-phosphate)-DNA + a 5'-end 5'-phospho- 2'-deoxyribonucleoside-DNA + H⁺

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

DOI no: 10.1074/jbc.RA118.006809

J Biol Chem 294:7194-7201 (2019)

PubMed id: 30885943

Molecular basis for the faithful replication of 5-methylcytosine and its oxidized forms by DNA polymerase β.

M.J.Howard, K.G.Foley, D.D.Shock, V.K.Batra, S.H.Wilson.

ABSTRACT

DNA methylation is an epigenetic mark that regulates gene expression in mammals. One method of methylation removal is through ten-eleven translocation-catalyzed oxidation and the base excision repair pathway. The iterative oxidation of 5-methylcytosine catalyzed by ten-eleven translocation enzymes produces three oxidized forms of cytosine: 5-hydroxmethylcytosine, 5-formylcytosine, and 5-carboxycytosine. The effect these modifications have on the efficiency and fidelity of the base excision repair pathway during the repair of opposing base damage, and in particular DNA polymerization, remains to be elucidated. Using kinetic assays, we show that the catalytic efficiency for the incorporation of dGTP catalyzed by human DNA polymerase β is not affected when 5-methylcytosine, 5-hydroxmethylcytosine, and 5-formylcytosine are in the DNA template. In contrast, the catalytic efficiency of dGTP insertion decreases ∼20-fold when 5-carboxycytosine is in the templating position, as compared with unmodified cytosine. However, DNA polymerase fidelity is unaltered when these modifications are in the templating position. Structural analysis reveals that the methyl, hydroxymethyl, and formyl modifications are easily accommodated within the polymerase active site. However, to accommodate the carboxyl modification, the phosphate backbone on the templating nucleotide shifts ∼2.5 Å to avoid a potential steric/repulsive clash. This altered conformation is stabilized by lysine 280, which makes a direct interaction with the carboxyl modification and the phosphate backbone of the templating strand. This work provides the molecular basis for the accommodation of epigenetic base modifications in a polymerase active site and suggests that these modifications are not mutagenically copied during base excision repair.