New technology illuminates protein interactions in living cells

Nov 09, 2007
New technology illuminates protein interactions in living cells
Schematic of fluorescent detector: When a target protein is folded correctly, "tags" come together so that the dye binds with high affinity and fluoresces brightly; misfolded proteins have low affinity for the dye. Credit: Schepartz/Nature Chemical Biology

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.

Proteins are commonly tagged using variants of the “green fluorescent protein” (GFP), 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,” said senior author Alanna Schepartz, the Milton Harris Professor of Chemistry, and Howard Hughes Medical Institute Professor at Yale. “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,” said 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.

Source: Yale University

Explore further: How can we avoid kelp beds turning into barren grounds?

add to favorites email to friend print save as pdf

Related Stories

Nail stem cells prove more versatile than press ons

Nov 21, 2014

There are plenty of body parts that don't grow back when you lose them. Nails are an exception, and a new study published in the Proceedings of the National Academy of Sciences (PNAS) reveals some of the r ...

Optical control of motor functions

Nov 10, 2014

MIT researchers have demonstrated a highly flexible neural probe made entirely of polymers that can both optically stimulate and record neural activity in a mouse spinal cord—a step toward developing prosthetic ...

Diet affects pesticide resistance in honey bees

Nov 03, 2014

Feeding honey bees a natural diet of pollen makes them significantly more resistant to pesticides than feeding them an artificial diet, according to a team of researchers, who also found that pesticide exposure ...

Cell division, minus the cells

Oct 30, 2014

(Phys.org) —The process of cell division is central to life. The last stage, when two daughter cells split from each other, has fascinated scientists since the dawn of cell biology in the Victorian era. ...

Recommended for you

Parasitic worm genomes: largest-ever dataset released

14 hours ago

The largest collection of helminth genomic data ever assembled has been published in the new, open-access WormBase-ParaSite. Developed jointly by EMBL-EBI and the Wellcome Trust Sanger Institute, this new ...

Male sex organ distinguishes 30 millipede species

14 hours ago

The unique shapes of male sex organs have helped describe thirty new millipede species from the Great Western Woodlands in the Goldfields, the largest area of relatively undisturbed Mediterranean climate ...

How can we avoid kelp beds turning into barren grounds?

18 hours ago

Urchins are marine invertebrates that mould the biological richness of marine grounds. However, an excessive proliferation of urchins may also have severe ecological consequences on marine grounds as they ...

User comments : 0

Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.