stearothermophilus Adenine DNA
Glycosylase Protein MutY
Elizabeth Bailey '12 and Jonathan Weil '11
The mutated base
has the potential to induce errors
in DNA replication. DNA polymerase misreads the oxoG and pairs it with
an adenine residue instead of the necessary cytosine residue (oxoG vs thymine).
uncorrected, this mispair can lead to permanent mutations following the
next round of replication. The adenine DNA glycosylase protein, MutY,
has the vital function of removing the mispaired A of the oxoG:A base
pair. A sequence specific glycosylase, MutY recognizes oxoG and cleaves
the mispaired adenine. MutY uses base flipping to twist the mispaired
adenine out of the DNA helix and into MutY’s active site.
Such adenine DNA glycosylases are found in both bacteria and humans
MutY and nMYH, respectivly). This tutorial presents the structure of
MutY from Bacillus
bound to DNA and inorganic
is composed of 2 distinct domains: the C-terminal
and the catalytic
C-terminal domain of
MutY contacts the oxoG
containing DNA strand and the catalytic domain. The catalytic domain is
composed of the
6-helix barrel module
4Fe-4S module .
C-terminal and catalytic domains are connected by an extended
that transverses the DNA
major groove near the lesion.
beneficial electrostatic interactions with the phosphate backbone
because it is rich in positively-charged lysines (Lys 228, 230, 231,
235). The contacts between the C-terminal domain and the catalytic
region allow MutY to encircle the DNA.
is completely extruded from the DNA helix, while the
modified guanine (oxoG) remains within the normal DNA structure.
adenine is held within a pocket in the catalytic domain of the protein.
with other members of
the structural superfamily base excision and
repair proteins, the catalytic domain in MutY contains a signature helix-hairpin-helix
element, followed by a Gly/Pro-rich
and a catalyticly essential Asn 144.
Interactions with 8-Oxoguanine
oxoG is found in the syn conformation when bound to adenine prior to
association with MutY.
glycosidic bond rotates
180° into the anti conformation in allow MutY to bind (OxoG syn vs. anti
conformation). This rotation is
thought to force the adenine to flip out of the
helix since the anti oxoG would clash sterically with the adenine. Tyr 88
is intercalated into the DNA on the 5' side of OxoG,
doubles the distance between OxoG and its 5' neighbor.
is inserted into the space vacated by the extruded adenine
and forms pi-stacking interactions with the 3'-neighboring
hydrogen bond to the phosphate 3' to the substrate adenine.
Gln 48 forms multiple direct and indirect stabilizing bonds to Tyr 88,
Ser 308, and the oxoG purine ring.
is also contacted
by the main
chain amide carbonyl of Gln 48.
further stabilized by
interactions with the side chain hydroxyl of Thr 49.
final pairs of
contacts are contributed by the main chain carbonyl of Leu 86 and the
main chain amide of Ser 308.
Ser 308 residue is
MutY’s discrimination between oxoguanine and guanine. If MutY
bound to a guanine, lost contacts would include the hydrogen bonding of
Ser 308’s N-H to oxoG’s O8, as well as the
extensive network between Ser 308, Tyr 88, and Gln 48.
Adenine Binding and Excision Reaction
removes the mispaired adenine by a dissociative SN1-like reaction mechanism
. The reaction has two distinct steps: breaking of the
gylcosidic bond and nucleophilic attack by water. MutY is able to
catalyze this reaction by stabilizing the reaction intermediates. Asn
144 is located near of the O4'
of the adenine
sugar and stabilizes the
charged ion intermediate in the base-excision step.
water below the C1'
of the adenine
sugar functions as the nucleophile
which reacts with the
charged intermediate at C1'
to form the final product.
molecule hydrogen bonds to the adenine N7, stabilizing the base leaving
hydrogen bond contacts with the Glu 43
43 functions as a
catalyst by accepting a proton then
transferring it to N7 via the water-bound intermediate, effectively
lowering the transition state energy for the base excision.
Fromme, J. C.; Banerjee, A.; Huang, S. J.;
and G. L Verdine. 2004.
Structural basis for removal of adenine mispaired with 8-oxoguanine by
MutY adenine DNA glycosylase. Nature
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