Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

Polarized microscopy technique shows new details of how proteins are arranged

18.04.2011
Whether you're talking about genes, or neurons, or the workings of a virus, at the most fundamental level, biology is a matter of proteins.

So understanding what protein complexes look like and how they operate is the key to figuring out what makes cells tick. By harnessing the unique properties of polarized light, Rockefeller scientists have now developed a new technique that can help deduce the orientation of specific proteins within the cell.

By turning their instruments toward the nuclear pore complex, a huge cluster of proteins that serves as a gateway to a cell's nucleus, the scientists say they have filled in the gaps left by other techniques and made important new discoveries about how the complex works.

"Our new technique allows us to measure how components of large protein complexes are arranged in relation to one another," says Sandy Simon, head of the Laboratory of Cellular Biophysics. "This has the potential to give us important new information about how the nuclear pore complex functions, but we believe it can also be applied to other multi-protein complexes such as those involved in DNA transcription, protein synthesis or viral replication."

Although researchers have spent years studying the workings of the nuclear pore complex, there is still much that has remained mysterious. One problem is that there is a "resolution gap" between the two techniques primarily used to visualize protein complexes. Electron microscopy can reveal the broad outlines of a large protein complex, but it can't show details. X-ray crystallography, meanwhile, can show minute detail but only of a small piece of the complex; it can't say how the individual pieces fit together. To further complicate matters, both techniques require fixed samples – while they can give you an idea of what something looks like at a moment in time, they can't tell you how its pieces might move.

The new technique was developed by Simon along with postdoc Alexa Mattheyses, graduate student Claire Atkinson and Martin Kampmann, a former a member of Günter Blobel's Laboratory of Cell Biology who is currently at the University of California, San Francisco. It takes advantage of the properties of polarized light to show how specific proteins are aligned in relation to one another. After genetically attaching fluorescent markers to individual components of the nuclear pore complex, the scientists replaced the cell's own copy of the gene that encodes the protein with the new form that has the fluorescent tag. Then, they used customized microscopes to measure the orientation of the waves of light the fluorescently tagged proteins emitted. By combining these measurements with known data about the structure of the complex, the scientists can confirm or deny the accuracy of previously suggested models.

"Our experimental approach to the structure is synergistic with other studies being conducted at Rockefeller, including analysis with X-ray crystallography in Günter's lab and electron microscopy and computer analysis in Mike Rout's lab," says Simon. "By utilizing multiple techniques, we are able to get a more precise picture of these complexes than has ever been possible before."

The scientists used the technique to study nuclear pore complexes in both budding yeast and human cells. In the case of the human cells, their new data shows that multiple copies of a key building block of the nuclear pore complex, the Y-shaped subcomplex, are arranged head-to-tail, rather than like fence posts, confirming a model proposed by Blobel in 2007.

"As a graduate student with Günter Blobel, I determined the three-dimensional structure of the Y-shaped subcomplex using electron microscopy," says Kampmann. "However, it was still a mystery how these 'Y's are arranged. The new technique we have developed reveals the orientation of building blocks in the cell, and we hope that it will eventually enable us to assemble individual crystal structures into a high-resolution map of the entire nuclear pore complex."

Eventually, the scientists say their technique could go even further. Because the proteins' fluorescence can be measured while the cells are still alive, it could give scientists new insights into how protein complexes react to varying environmental conditions, and how their configurations change over time.

"What happens when other proteins pass through the nuclear pore? Does the orientation of the nucleoporins change? With this technique, can find out not only what the pore looks like when it's sitting still, but what happens to it when it's active," Simon says. Their first characterization of the dynamics of the nuclear pore proteins was published recently in The Biophysical Journal.

Zach Veilleux | EurekAlert!
Further information:
http://www.rockefeller.edu

More articles from Life Sciences:

nachricht Toward a 'smart' patch that automatically delivers insulin when needed
18.01.2017 | American Chemical Society

nachricht 127 at one blow...
18.01.2017 | Stiftung Zoologisches Forschungsmuseum Alexander Koenig, Leibniz-Institut für Biodiversität der Tiere

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: How gut bacteria can make us ill

HZI researchers decipher infection mechanisms of Yersinia and immune responses of the host

Yersiniae cause severe intestinal infections. Studies using Yersinia pseudotuberculosis as a model organism aim to elucidate the infection mechanisms of these...

Im Focus: Interfacial Superconductivity: Magnetic and superconducting order revealed simultaneously

Researchers from the University of Hamburg in Germany, in collaboration with colleagues from the University of Aarhus in Denmark, have synthesized a new superconducting material by growing a few layers of an antiferromagnetic transition-metal chalcogenide on a bismuth-based topological insulator, both being non-superconducting materials.

While superconductivity and magnetism are generally believed to be mutually exclusive, surprisingly, in this new material, superconducting correlations...

Im Focus: Studying fundamental particles in materials

Laser-driving of semimetals allows creating novel quasiparticle states within condensed matter systems and switching between different states on ultrafast time scales

Studying properties of fundamental particles in condensed matter systems is a promising approach to quantum field theory. Quasiparticles offer the opportunity...

Im Focus: Designing Architecture with Solar Building Envelopes

Among the general public, solar thermal energy is currently associated with dark blue, rectangular collectors on building roofs. Technologies are needed for aesthetically high quality architecture which offer the architect more room for manoeuvre when it comes to low- and plus-energy buildings. With the “ArKol” project, researchers at Fraunhofer ISE together with partners are currently developing two façade collectors for solar thermal energy generation, which permit a high degree of design flexibility: a strip collector for opaque façade sections and a solar thermal blind for transparent sections. The current state of the two developments will be presented at the BAU 2017 trade fair.

As part of the “ArKol – development of architecturally highly integrated façade collectors with heat pipes” project, Fraunhofer ISE together with its partners...

Im Focus: How to inflate a hardened concrete shell with a weight of 80 t

At TU Wien, an alternative for resource intensive formwork for the construction of concrete domes was developed. It is now used in a test dome for the Austrian Federal Railways Infrastructure (ÖBB Infrastruktur).

Concrete shells are efficient structures, but not very resource efficient. The formwork for the construction of concrete domes alone requires a high amount of...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

12V, 48V, high-voltage – trends in E/E automotive architecture

10.01.2017 | Event News

2nd Conference on Non-Textual Information on 10 and 11 May 2017 in Hannover

09.01.2017 | Event News

Nothing will happen without batteries making it happen!

05.01.2017 | Event News

 
Latest News

A big nano boost for solar cells

18.01.2017 | Power and Electrical Engineering

Glass's off-kilter harmonies

18.01.2017 | Materials Sciences

Toward a 'smart' patch that automatically delivers insulin when needed

18.01.2017 | Life Sciences

VideoLinks
B2B-VideoLinks
More VideoLinks >>>