Monday, August 2, 2010

Berkeley Lab news: A cellular housekeeper, and potential target of obesity drugs, caught in action

A cellular housekeeper, and potential target of obesity drugs, caught in
action

online news release with images:
http://newscenter.lbl.gov/news-releases/2010/08/01/cellular-housekeeper/

contact: Dan Krotz, dakrotz@lbl.gov, (510) 486-4019

Scientists from the U.S. Department of Energy's Lawrence Berkeley
National Laboratory have obtained the closest look yet of how a
gargantuan molecular machine breaks down unwanted proteins in cells, a
critical housekeeping chore that helps prevent diseases such as cancer.

They pieced together the molecular-scale changes the machine undergoes
as it springs into action, ready to snip apart a protein.

Their work provides valuable clues as to how the molecular machine, a
giant enzyme called tripeptidyl peptidase II, keeps cells tidy and
disease free. It could also inform the development of obesity-fighting
drugs. A closely related enzyme in the brain can cause people to feel
hungry even after they eat a hearty meal.

"We can now better understand how this very important enzyme carries out
its work, which has not been described at a molecular scale until now,"
says Bing Jap, a biophysicist in Berkeley Lab's Life Sciences Division.
He led the research with scientists from the University of California at
Berkeley and Germany's Max Planck Institute of Biochemistry.

The scientists report their research August 1 in an advance online
publication of the journal Nature Structural & Molecular Biology.

Tripeptidyl peptidase II is found in all eukaryotic cells, which are
cells that a have membrane-bound nucleus. Eukaryotic cells make up
plants and animals. The enzyme's chief duty is to support the pathway
that ensures that cells remain healthy and clutter free by breaking down
proteins that are misfolded or have outlived their usefulness.

It's not always so helpful, however. A variation of the enzyme in the
brain degrades a hormone that makes people feel satiated after a meal.
When this hormone becomes unavailable, a person can eat and eat without
feeling full, which can lead to obesity.

Tripeptidyl peptidase II is also the largest protein-degrading enzyme,
or protease, in eukaryotic cells. It's more than 100 times larger than
most other proteases.

Scientists don't know how this behemoth of an enzyme targets and
degrades specific proteins — but it's good that the enzyme is so
selective. If it degraded every protein it comes across, the cell would
quickly die.

"We want to know how it's regulated, how it selects proteins to degrade,
and how it cuts them apart," says Jap.

To help answer these questions, his team determined the changes the
molecular machine undergoes as it readies itself for action. Using x-ray
crystallography, they obtained an atomic-scale resolution structure of
the molecular machine in its inactive state. This work was conducted at
Berkeley Lab's Advanced Light Source, a national user facility that
generates intense x-rays to probe the fundamental properties of substances.

They also developed a lower-resolution, three-dimensional map of the
molecular machine in its activated state, meaning it's poised to snip
apart a protein. This structure was determined using cryo-electron
microscopy.

They then merged these two structures together, one dormant and the
other ready to pounce on a protein.

"When we dock these structures, we can begin to ascertain the changes
the enzyme undergoes as it transitions from an inactive to an active
state," says Peter Walian, a scientist in Berkeley Lab's Life Sciences
Division who also contributed to the research.

This first molecular-scale vantage of the enzyme in action offers
insights into how it works. For example, the scientists found that only
very small proteins can fit in the chamber the enzyme uses to break down
proteins.

"This sheds light on how the enzyme targets specific proteins," says Jap.

They also learned more about how the enzyme uses a molecular ruler to
mince proteins into pieces that only span three residues.

"This work is yielding valuable clues as to how the giant enzyme carries
out very fundamental biological processes, with more insights to come,"
says Jap. "The obesity-related hormone is one of many interesting
targets of the protease. There are likely other proteins and peptides,
yet to be discovered, that are processed by this protease."

The research was supported by the National Institute of General Medical
Sciences of the National Institutes of Health. The Advanced Light Source
is supported by the Department of Energy's Office of Science.

Lawrence Berkeley National Laboratory provides solutions to the world's
most urgent scientific challenges including clean energy, climate
change, human health, and a better understanding of matter and force in
the universe. It is a world leader in improving our lives and knowledge
of the world around us through innovative science, advanced computing,
and technology that makes a difference. Berkeley Lab is a U.S.
Department of Energy (DOE) national laboratory managed by the University
of California for the DOE Office of Science. This content is solely the
responsibility of Lawrence Berkeley National Laboratory. Visit our
website: http://www.lbl.gov/

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