protein interactions in living cells
While fluorescence has long been used to tag biological molecules, a new
technology developed at Yale allows researchers to use tiny fluorescent probes
to rapidly detect and identify protein interactions within living cells while
avoiding the biological disruption of existing methods, according to a report
in Nature Chemical Biology. — By Janet Rettig Emanuel
T H I S
SCHOOL OF MEDICINE NEWS
Proteins are commonly tagged using variants of the “green fluorescent protein,” but
these proteins are very large and are often toxic to live cells. They also tend
to aggregate, making them difficult to work with and monitor. This new methodology
uses the fluorescence emitted by a small molecule, rather than a large protein.
It gives researchers a less disruptive way to capture images of the intricate
contacts between folded regions of an individual protein or the partnerships
between proteins in a live cell.
“Our approach bypasses many of the problems associated with fluorescent
proteins, so that we can image protein interactions in living cells,” says
senior author Alanna Schepartz, the Milton Harris Professor of Chemistry and
a Howard Hughes Medical Institute investigator. “Using these molecules
we can differentiate alternative or misfolded proteins from those that are folded
correctly and also detect protein partnerships in live cells.”
Each protein is a three-dimensional structure created by “folding” its
linear chain of amino acids. Usually only one shape “works” for each
protein. The particular shape a protein takes depends on its amino acids and
on other processes within the cell.
Schepartz and her team devised their new tagging system using small molecules,
called “profluorescent” biarsenal dyes. These molecules easily enter
cells and become fluorescent when they bind to a specific amino acid tag sequence
within a protein. While these compounds have been used for about a decade to
bind single proteins, this is the first time they have been used to identify
interactions between proteins.
The researchers’ strategy was to split the amino acid tag for the dye into
two pieces, locating each piece of the tag far apart in the chain of a protein
they genetically engineered and expressed in the cells. Then they monitored cells
exposed to the dye. Where the protein folded correctly, the two parts of the
tag came together and the fluorescent compound bound and lit up. There was no
signal unless the protein folded normally.
“This method of detection can provide important insights into how proteins
choose their partners within the cell — choices that may be very different
from those made in a test tube,” said Schepartz. She emphasizes that this
technology does not monitor the process of protein folding but, rather, “sees” the
protein conformations that exist at a given time.
“In theory, our technique could be used to target and selectively inactivate
specific protein complexes in the cell, as therapy, or to visualize conformations
at very high resolution for diagnostic purposes,” says Schepartz. She speculates
that the technology could be applied to detection strategies that identify protein
misfolding in neurodegenerative diseases like Alzheimer’s or Parkinson’s.
Other authors on the paper are Nathan W. Luedtke, Rachel J. Dexter and Daniel
B. Fried from the Schepartz lab at Yale. Funding from the Howard Hughes Medical
Institute and the National Institutes of Health supported the research.
W E E K ' SS T O R I E S
Restoration of iconic Rudolph building is key step in . . . Arts Complex
Three faculty win nation’s highest award for beginning researchers
Bulldogs, Crimsons both bringing undefeated Ivy records . . .
A grateful nation
New technology allows view of protein interactions in living cells
Monkeys and children share adults’ tendency to rationalize choices
Noted composer Benjamin Lees donates his archive to Yale library
Museum joins with area public schools to promote ‘visual literacy’
A conversation in China leads to successful research collaboration
Junior faculty earn second terms in endowed posts
Study: Simple measures could prevent spread of . . . tuberculosis
Erol Fikrig, specialist in vector-borne diseases . . .
Physical and emotion stress linked to aortic rupture
Groundbreaking scientist joins Yale Stem Cell Center
Team develops blood test for deadly aneurysm disease
Workshop will showcase new Positron Emission Tomography Center
Four decades of readers in Yale libraries are featured in exhibition
Video installations by Ori Gersht on view at British Art Center
Reception will celebrate United Way donors as campaign continues
Yale Books in Briefs
Benefit event to feature noted neurosurgeon
Workshop to feature Ohio State law professor
Reminder: Open enrollment period ends Nov. 18
Campus Notes
Bulletin Home|Visiting on Campus|Calendar of Events|In the News
Bulletin Board|Classified Ads|Search Archives|Deadlines
Bulletin Staff|Public Affairs|News Releases|
E-Mail Us|Yale Home