Scientists Uncover Critical Step in DNA Mutation
Atlanta (August 23, 2006) — Scientists at the Georgia Institute of Technology have made an important step toward solving a critical puzzle relating to a chemical reaction that leads to DNA mutation, which underlies many forms of cancer. The research, which uncovers knowledge that could be critical to the development of strategies for cancer prevention and treatment, appears in the August 2006 edition (Volume 128, issue 33) of the Journal of the American Chemical Society.
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Overview showing the position of
the actors in the reaction . For the full image
click below. For the full caption see bottom. |
The process that gives rise to mutations in DNA, or
mutagenesis, is a complex one involving a series of chemical
reactions, which are not completely understood. A free radical,
a stable neutral atom or a chemical group containing at least
one unpaired electron, can scavenge an electron from DNA in a
process known as oxidation, creating a hole in place of the
scavenged electron. Such oxidation events can be caused by
natural processes occurring in the body, or by ionizing
radiation. It’s well known that the ionization hole can travel
long distances of up to 20 nanometers along the base pairs that
form the rungs of the DNA ladder (discussed by Landman, Schuster
and their collaborators in a 2001 Science article, volume 294,
page 567). It is also well known that the hole tends to settle
longer at spots in the DNA where two guanines (G) are located
next to each other.
It’s the next step that has eluded DNA researchers for decades -
somehow the hole in the ionized DNA reacts with water. This
critical step is the first in a series that brings about a
change in the DNA molecule – one that evades the body’s proof
reading mechanism and leaves the altered DNA coding for the
wrong proteins. When the wrong proteins are produced, it can
lead through a complicated chain of events to an abnormally high
rate of cell division – the result is cancer.
"We set out to explore the elementary processes that lead to
mutagenesis and eventually cancer,” said Uzi Landman, director
of the Center for Computational Materials Science and Regents’
professor and Callaway chair of physics at Georgia Tech.
"Until now, the mechanism by which water reacts with the guanine
of the ionized DNA remained a puzzle. Through our
first-principles, computer-based quantum mechanical theoretical
modeling, coupled with theory-driven laboratory experiments, we
have gained important insights into a critical step in a
reaction that can have far reaching health consequences,” he
said.
Once the hole is settled on the two guanine bases, water
molecules react with one of the bases at a location called the
8-th carbon site (C8). This reaction converts it into 8-oxo-7,
8-dihyrdroguanine (8-Oxo-G). But, this reaction requires more
energy than seems to be available because, formally, it requires
that a water molecule (H2O) split apart into a proton (H+) and a
hydroxyl anion (OH-). This large energy requirement has puzzled
scientists for a long time. Now the research team, led by
Landman and Gary Schuster, provost-designate of Georgia Tech,
professor of Chemistry and dean of the College of Sciences, has
uncovered how the reaction occurs.
 |
Detail view of the reaction of
water with the guanine radical cation. For the full
image click below. For the full caption see bottom. |
Here’s what they found: A sodium counter-ion (Na+) diffusing
in the hydration environment of the DNA molecule wanders into
the major groove of the DNA double helical ladder. When the Na+
comes close to the hole created by the missing electron, its
positive charge promotes the C8 carbon atom of the guanine to
bond with a water molecule. In a concerted motion, the oxygen
atom of the water molecule with one of its hydrogen atoms
attaches to the C8 carbon atom. At the same time, the other
proton of the water molecule connects the oxygen atom to that of
a neighboring water molecule. This hydrogen bond elongates,
leading to the formation of a transition state complex involving
the two neighboring water molecules. The complex breaks up,
transferring one of its protons (a positively charged hydrogen
atom) to the neighboring water molecule, making a hydronium ion
H3O+. This leaves the guanine neutral, with the rest of the
first water molecule attached to it and prepares it for the rest
of the already-known steps to making 8-Oxo-G.
Because it is positively charged, the H3O+ binds to the adjacent
negatively charged phosphate, (PO4-) that is part of the
backbone of the DNA molecule, to complete this step in the
reaction.
The phosphate is crucial to the reaction because it acts as a
sink that holds one of the reaction products, (H3O+) together
with the other product (the guanine base with an attached OH- at
the C8 location). According to quantum simulations, the energy
barrier leading to formation of the transition state complex,
and thus the required energy for this reaction step to occur, is
0.7 electron-volts (eV) – well below the energy required for
dissociation of a water molecule immersed in a water
environment. Obviously, without the presence of neighboring
water molecules, the above reaction mechanism involving transfer
of the proton to the neighboring phosphate group through the
hydronium shuttle, does not occur and no products are generated.
The simulations unveiled that the Na+ plays a key role in
promoting the reaction. To test this theoretical prediction in
the laboratory, the team substituted the negatively charged PO4-
near the reaction site with a phosphonate (PO3CH3) group.
Because phosphonate is neutral, it doesn’t attract the Na+.
Without the Na+ to promote the reaction of the ionized DNA at
the C8 carbon atom of the guanine base, the reaction becomes
less likely. Furthermore, even if a reaction occurs and a H3O+
forms, it does not get attached to the neutral phosphonate, and
consequently the reaction does not come to completion, and very
little, or no 8-oxo-G is formed.
"The complexity of this reaction is an intrinsic part of the
chemical process that we investigated, because it occurs only
under very specific conditions requiring a complex choreography
from its players, I believe that this complexity is part of
nature’s control mechanism,” said Landman. “Perhaps such
inherent complexity guards us from harmful mutagenetic events
occurring more frequently, and it is possible that similar
principles may hold in other important processes of biological
relevance.”
"This type of research requires the development of new modeling
strategies and significant computational power. It also needed
indispensable complementary and supplementary laboratory
experiments. We were very fortunate to have this combination in
our research team,” he said.
The authors of the paper published in the Journal of the
American Chemical Society are:
Robert N. Barnett, Angelo Bongiorno, Charles L. Cleveland,
Abraham Joy, Uzi Landman and Gary B. Schuster from the Schools
of Physics and Chemistry and Biochemistry at the Georgia
Institute of Technology.
Image one caption:
The ionized 14-base pair oligomer of B-DNA
[d(5'-AAGGAAGGAAGGAA-3')]/[d(3'-TTCCTTCCTTCCTT-5')], modeled in
this study through a hybrid quantum/classical simulation method.
The bold letters GG/CC in the middle of the sequence denote the
region treated quantum mechanically (QM), where the ionization
hole reside. The hole density, depicted in light blue, is
superimposed.on the QM region. In the background we show the
water environment surrounding the DNA molecule. In the QM region
the color assignments are as follows: P yellow, C green, N blue,
O (base) red, O (phosphate) red, O (H2O) orange, H (H2O) small
blue spheres, Na purple. Note that most of the counter ions are
located in the vicinity of the phosphate groups, with one of the
counter ions in the QM region residing in them major groove.
Image two caption:
Steps of the reaction of water with the guanine radical cation
in DNA. Only a small part of the system is shown, focusing on
the reaction site (C8) and the immediate water molecules, with
the oxygen of the attacking H2O molecule denoted as O1. A) step
1: the relaxed ionized configuration. (B) Step 2: the approach
of an H2O molecule to C8 of the guanin base is accompanied by
elongation of the dO1-H11 bond of the molecule along the axis
connecting the oxygen to that of a neighboring water molecule,
and the activated formation of a transition state complex (with
a barrier of about 0.7 eV). (C) Step 3: further evolution of the
reaction leads to breakup of the complex, with formation of a
hydroxylated G radical (8-OH-3'-G.) and a concomitant proton
shuttle process, ending with the attachment of an asymmetric
hydronium group to the phosphate. The color assignments are as
follows: P yellow, C gray, N blue, O red, H white. The reaction
site is labeled C8 of the 3'-G, and the oxygen of the attacking
H2O molecule is O1.
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(Source: Tech News Release)
