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Lock and key
The work is motivated by a desire to map the
three-dimensional requirements of biological receptors on cell
surfaces. Typically, receptors bind small molecules through a lock and
key mechanism where the molecule is the key and the receptor the lock.
The nature and shape of molecules that serve as keys tells about the
binding requirements of the receptor. Traditionally, probing a
receptor this way has been done by making a library of molecules,
treating it with the receptor, washing away any excess receptor that
has not found a key, and then treating the bound receptors with an
antibody that recognizes the receptor and is tagged to a fluorescent
label. The washing step risks removing a bound receptor if it does not
bind the molecular key strongly enough. But, with an electrochemically
addressable computer chip, provided in great abundance by one of his
sponsor's, CombiMatrix in Seattle, Moeller saw a way of probing the
binding of a library with a receptor without the need for washing by
putting each member of the molecular library by an electrode that can
then be used to monitor its behavior.
The electrochemically addressable chips being used
represent a new environment for synthetic organic chemistry, changing
the way chemists and biomedical researchers make molecules, build
molecular libraries and understand the mechanisms by which molecules
bind to receptor sites.
"We believe we can move most of modern synthetic
organic chemistry to this electrochemically addressable chip. In this
way, a wide variety of molecules can be generated and then probed for
their biological behavior in real-time," said Moeller. "It's a tool,
still being developed, to map receptors. We're right at the cusp of
things."
Moeller published on the technology in a recent
article in the Journal of the American Chemical Society, Vol. 28
16020, 2006.
The Great Wall of China syndrome
Moeller said that the standard problem for
synthetic organic chemists has typically been a structural one - how
do we build molecules having novel structures?
"We've worked very hard to develop new chemical
reactions that allow us to make new structures that are either
difficult or impossible to make with the synthetic tools available,"
he said. "But now for the first time it's not just a matter of
structure, but rather a matter of location and scale - a logistical
problem."
He offered the Great Wall of China as an example of
a classic logistics problem - the structure itself was simple, but
getting the tools and manpower necessary for the task to the remote
regions it was built in on the scale necessary was "a logistical
nightmare."
Getting 12,000 electrodes per square centimeter to
selectively do your chemical bidding is Moeller's logistical nightmare.
How do you get chemical reactions to happen at just one of the
electrodes? Here's how he and his colleagues addressed it.
Scientists at CombiMatrix initially pioneered an
approach for covering the chip with a polymer, attaching a substrate
to the polymer right above the electrode, and then using the electrode
to initiate a chemical reaction that modified the substrate and
converted it into a product. Putting a confining agent into the
solution destroys the reagent made, keeping it from going to another
electrode. This was done for the generation of acids and bases and
applied to the synthesis of DNA and peptide polymers. But could this
approach be used to make the more difficult to synthesize shaped
molecules needed for mapping a receptor? This would require a diverse
array of reactions to be compatible with the chips, particularly
reactions that capitalized on modern, state-of-the-art metal-based
reagents.
Working with his students, Moeller started by
running a well-known electrochemical Wacker oxidation reaction, which
converts an olefin into a ketone using palladium(II) on selected
electrodes on the chip. In order to do this, the palladium(II) reagent
needed at a selected electrode was actually made at the desired site
using the electrode. This allowed for the desired reaction at only
electrodes that were turned on. To keep the palladium(II) from going
to a neighboring electrode a second reagent was added to the solution
above the chip that destroyed the palladium(II) reagent. In this way,
nothing happened at electrodes that were not being used.
Reversing the polarity of things
"The chip is an ongoing battle between active
reagent and inactive reagent," Moeller said. "The active reagent is
fine - it does the chemistry I want it to do - but as soon as it gets
away from the turned-on electrode the reaction that inactivates it
needs to take over."
With an overall strategy in place, Moeller and his
collaborators are working to expand the scope of reactions that can be
done. For example, they have carried out the oxidation of an alcohol
to form a carbonyl and then used the carbonyl to put a dye down. They
placed a green dye at alternating electrodes in a checkerboard pattern,
and then used the alternate set of electrodes to put down a red dye.
The result was a red and green chip, Washington University's colors.
This, Moeller said, is a striking image of the technique's ability to
do chemistry at specific electrodes. "What electrochemistry does best
is to reverse the polarity of things," Moeller said. "It takes things
that are electron-poor and makes them electron-rich and vice versa; it
can take an oxidant and turn it into a reductant; a reductant into an
oxidant; a base into an acid and an acid into a base; and so on. In
the end, almost any chemical reagent can be made at an electrode and
used on the chips."
Moeller said that , along with emphasizing new
reactions, he and his collaborators are working to better identify the
molecules they make on the chips, and perfect the signaling techniques
used to monitor molecules on the chips. He praised the time-of-flight
secondary-ion-mass spectrometry work of his Washington University
colleague Amy Walker, Ph.D., assistant professor of chemistry, in the
monitoring of his system. |
