PDBsum entry 5ugp

Transferase/DNA

PDB id: 5ugp

Contents

Protein chain

323 a.a.

DNA/RNA

Ligands

8CP

Metals

_MG ×2

Waters ×318

PDB id:

5ugp

Name:

Transferase/DNA

Title:

DNA polymerase beta complex with a 1nt gap and dcmppnp

Structure:

DNA polymerase beta. Chain: a. Engineered: yes. DNA (5'-d(p Gp Tp Cp Gp G)-3'). Chain: d. Engineered: yes. DNA (5'-d( Gp Cp Tp Gp Ap Tp Gp Cp Gp C)-3'). Chain: p. Engineered: yes.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: polb. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Synthetic construct. Organism_taxid: 32630.

Resolution:

1.96Å R-factor: 0.185 R-free: 0.233

Authors:

B.D.Freudenthal,S.H.Wilson,W.A.Beard

Key ref:

D.D.Shock et al. (2017). Modulating the DNA polymerase β reaction equilibrium to dissect the reverse reaction. Nat Chem Biol, 13, 1074-1080. PubMed id: 28759020

Date:

09-Jan-17 Release date: 26-Jul-17

Protein chain

P06746  (DPOLB_HUMAN) -  DNA polymerase beta from Homo sapiens

Seq: 335 a.a.

Struc: 323 a.a.

DNA/RNA chains

G-T-C-G-G 5 bases

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

C-C-G-A-C-G-G-C-G-C-A-T-C-A-G-C 16 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

Nat Chem Biol 13:1074-1080 (2017)

PubMed id: 28759020

Modulating the DNA polymerase β reaction equilibrium to dissect the reverse reaction.

D.D.Shock, B.D.Freudenthal, W.A.Beard, S.H.Wilson.

ABSTRACT

DNA polymerases catalyze efficient and high-fidelity DNA synthesis. While this reaction favors nucleotide incorporation, polymerases also catalyze a reverse reaction, pyrophosphorolysis, that removes the DNA primer terminus and generates deoxynucleoside triphosphates. Because pyrophosphorolysis can influence polymerase fidelity and sensitivity to chain-terminating nucleosides, we analyzed pyrophosphorolysis with human DNA polymerase β and found the reaction to be inefficient. The lack of a thio-elemental effect indicated that this reaction was limited by a nonchemical step. Use of a pyrophosphate analog, in which the bridging oxygen is replaced with an imido group (PNP), increased the rate of the reverse reaction and displayed a large thio-elemental effect, indicating that chemistry was now rate determining. Time-lapse crystallography with PNP captured structures consistent with a chemical equilibrium favoring the reverse reaction. These results highlight the importance of the bridging atom between the β- and γ-phosphates of the incoming nucleotide in reaction chemistry, enzyme conformational changes, and overall reaction equilibrium.