such as "Introduction", "Conclusion"..etc
at TUM, the Technische Universitaet Muenchen, have published the
results of single-molecule experiments that bring a higher-resolution
tool to the study of protein folding. How proteins arrive at the
three-dimensional shapes that determine their essential functions – or
cause grave diseases when folding goes wrong – is considered one of the
most important and least understood questions in the biological and
medical sciences. Folding itself follows a path determined by its
energy landscape, a complex property described in unprecedented detail
by the TUM researchers. In this week's issue of the Proceedings of the National Academy of Sciences (USA),
they report taking hold of a single, zipper-like protein molecule and
mapping changes in its energy landscape during folding and unfolding.
studies, including atomic force microscopy experiments by the same
Munich laboratory, have gone a long way toward characterizing energy
thresholds or barriers that stand between a protein's unfolded and
folded states. Detailed observations of the quick transition from one
state to the other have remained elusive. The results published this
week open the door to higher-resolution, direct measurements. Better
characterization of the folding process is seen as a vital link in
understanding the chain of events leading from DNA coding for a protein
to that protein's biological function. Another motivation for research
in this field is the search for new drugs and therapies, because
malfunctions in protein folding are implicated in a number of serious
diseases – including diabetes, cancer, cystic fibrosis, prion diseases,
This is the latest in a long series of
single-molecule biophysical experiments carried out by Professor
Matthias Rief and colleagues in the TUM Department of Physics.
Co-authors Christof Gebhardt and Thomas Bornschloegl are members of
Rief's lab; Gebhardt also is a member of the Munich Center for
Integrated Protein Science.
As a model system for studying
real-time protein folding dynamics, the TUM scientists chose a
so-called leucine zipper found in yeast. It offers, as proteins go, a
relatively simple "coiled coil" structure and zipper-like folding
action: Picture two amino acid strings side by side, joined at the
bottom, open at the top, and made essentially to zip together.
researchers extended this structure so that they could make independent
measurements at the top, bottom, and middle parts of the zipper. They
took hold of the free ends at the top of the zipper with handles made
of double-stranded DNA. These DNA handles in turn were attached to tiny
beads that could be directly manipulated by "optical tweezers" – a tool
based on the ability of laser beams with a certain kind of profile to
pin down nanoscale objects. One end of the protein molecule was held
fixed, and the other was held under tension but with some freedom to
move, so that folding dynamics could be measured directly, in real
time, as the protein zipped and unzipped. This arrangement enabled
measurements with high resolution in both space and time.
I consider the major improvement is that the new experiments allow the
observation of thousands of transitions between the folded and the
unfolded state," Rief said. "This enables us to detect not only the
folded and unfolded states but also, directly, the excursions of the
large energy barriers separating those states. This has previously been
impossible, and it now allows direct insight into the precise energy
profile of this barrier."
Technische Universitaet Muenchen
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