Summary information and primary citation
- PDB-id
-
9icu;
SNAP-derived features in text and
JSON formats
- Class
- transferase-DNA
- Method
- X-ray (2.9 Å)
- Summary
- DNA polymerase beta (pol b) (e.c.2.7.7.7) complexed
with six base pairs of DNA; soaked in the presence of dttp
(1 millimolar) and mncl2 (5 millimolar)
- Reference
-
Pelletier H, Sawaya MR, Wolfle W, Wilson SH, Kraut J
(1996): "A
structural basis for metal ion mutagenicity and
nucleotide selectivity in human DNA polymerase beta."
Biochemistry, 35, 12762-12777.
doi: 10.1021/bi9529566.
- Abstract
- When crystals of human DNA polymerase beta (pol beta)
complexed with DNA [Pelletier, H., Sawaya, M. R., Wolfle,
W., Wilson, S. H., & Kraut, J. (1996) Biochemistry 35,
12742-12761] are soaked in the presence of dATP and Mn2+,
X-ray structural analysis shows that nucleotidyl transfer
to the primer 3'-OH takes place directly in the crystals,
even though the DNA is blunt-ended at the active site.
Under similar crystal-soaking conditions, there is no
evidence for a reaction when Mn2+ is replaced by Mg2+,
which is thought to be the divalent metal ion utilized by
most polymerases in vivo. These results suggest that one
way Mn2+ may manifest its mutagenic effect on polymerases
is by promoting greater reactivity than Mg2+ at the
catalytic site, thereby allowing the nucleotidyl transfer
reaction to take place with little or no regard to
instructions from a template. Non-template-directed
nucleotidyl transfer is also observed when pol beta-DNA
cocrystals are soaked in the presence of dATP and Zn2+, but
the reaction products differ in that the sugar moiety of
the incorporated nucleotide appears distorted or otherwise
cleaved, in agreement with reports that Zn2+ may act as a
polymerase inhibitor rather than as a mutagen [Sirover, M.
A., & Loeb, L. A. (1976) Science 194, 1434-1436]. Although
no reaction is observed when crystals are soaked in the
presence of dATP and other metal ions such as Ca2+, Co2+,
Cr3+, or Ni2+, X-ray structural analyses show that these
metal ions coordinate the triphosphate moiety of the
nucleotide in a manner that differs from that observed with
Mg2+. In addition, all metal ions tested, with the
exception of Mg2+, promote a change in the side-chain
position of aspartic acid 192, which is one of three highly
conserved active-site carboxylate residues. Soaking
experiments with nucleotides other than dATP (namely, dCTP,
dGTP, dTTP, ATP, ddATP, ddCTP, AZT-TP, and dATP alpha S)
reveal a non-base-specific binding site on pol beta for the
triphosphate and sugar moieties of a nucleotide, suggesting
a possible mechanism for nucleotide selectivity whereby
triphosphate-sugar binding precedes a check for correct
base pairing with the template.
