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Typically, analytical chemists use mass spectrometry - a technique
that accurately weighs molecules or fragments of molecules - to
analyze proteins. In this process, proteins are introduced into the
mass spectrometer and fragmented by heating until the weakest bonds
break. "It's the 'shake-it-til-it-breaks' approach," Hakansson said.
Together, the masses of the various fragments provide a sort of
fingerprint that reveals the genetic blueprint from which the protein
was built - information that helps researchers confirm the protein's
identity. This works fine as long as the protein has not been modified
after it was produced. But if other chemical groups such as phosphates,
sulfates or sugars have been added, the identification method breaks
down.
"If sugars are attached, for instance, the weakest bonds are not the
bonds that hold the protein together; they're the bonds between the
sugars," Hakansson said. When those bonds break, the resulting
fragments don't give accurate information about either the protein's
identity or the exact type and position of sugars present.
To get around that problem, researchers have used a process called
electron capture dissociation (ECD) instead of the usual "shake-it-til-it
breaks" method to fragment proteins. But that method requires the
presence of at least two positive charges, which can be difficult to
accomplish with acidic molecules, such as proteins with sulfate or
phosphate groups attached.
Hakansson's group has been exploring the use of metals such as calcium
and iron to carry the necessary positive charges. In a series of
recently published papers, they first showed that their method can be
used to selectively cleave different bonds and then demonstrated that
it can be used to identify sulfate-laden proteins and to pinpoint the
location of the sulfate groups on them.
In the latest research, they extended the technique to sugars, an even
more challenging task.
"Sugars are not like other biomolecules," Hakansson said. "They're
linked rings with lots of branches, like trees. If you cut off a
branch, you don't know which part of the tree it came from." The trick
is to make breaks that cut across the ring structures, rather lopping
off branches. By using metals as charge carriers, the researchers were
able to do just that, yielding valuable structural information.
In a project that continues to build on this line of work, Hakansson
is collaborating with U-M Health System cancer surgeon Diane Simeone
to investigate sugars attached to proteins in the membranes of
pancreatic cancer cells.
"The work is in very early stages, but we hope that by measuring
unique sugars it may be possible to develop diagnostic tools or
therapeutic agents to specifically target them," Hakansson said.
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