Why spiders don't drop off of their threads

It has five times the tensile strength of steel and is stronger then even the best currently available synthetic fibers: Spider thread. German scientists of the Technische Universitaet Muenchen and the Universitaet Bayreuth have now succeeded in unveiling a further secret of silk proteins and the mechanism that imparts spider silk with its strength. They have published the results of their work in the professional journal Angewandte Chemie.

"The strength of spider dragline silk exceeds that of any material produced in laboratories, by far. All attempts to manufacture threads of similar strength have failed thus far," explains Professor Horst Kessler, Carl von Linde Professor at the Institute for Advanced Study at the TU Muenchen (TUM-IAS). In collaboration with the workgroup of Prof. Thomas Scheibel, who was a researcher at the TU Muenchen until 2007 and who now holds a chair of the Institute of Biomaterials at the Universitaet Bayreuth, Professor Kessler's team has been researching for years to unveil the secret of spider silk.

How do spiders manage to first store the silk proteins in the silk gland and to then assemble them in the spinning passage in a split second to form threads with these extraordinary characteristics? And what exactly gives the threads their tremendous tensile strength? Scientists have now come one step closer to answering these key questions for the production of artificial spider silk.

Spider threads consist of long chains of thousands of repeating sequences of protein molecules. These silk proteins are stored in the silk gland in a highly concentrated form until they are needed. The long chains with their repeating sequences of protein molecules are initially unordered and must not get too close to each other as they would immediately clump up. Only in the spinning passage, just before being used, are the threads oriented parallel to each other and form so-called micro crystallites that are, in turn, assembled to stable threads with cross links.

During the last year, the scientists in Kessler's and Scheibel's team investigated the common garden spider ("cross spider") to discover the mechanism behind the transition from individual spider silk molecules to connected treads: The individual spider silk proteins are first stored in the silk gland in small drops called micelles.

The scientists identified the regulating element that is responsible for assembling a strong thread from the individual parts. It is the so-called C terminal domain of the silk protein. It prevents the formation of threads in the silk duct with its strong salt concentrations. In the spinning passage, however, where the salt concentration is low and sheer forces are abundant, this domain becomes instable and "sticky." This causes the chains to overlap and a strong spider silk thread is formed. The discovery of the significance of this relatively small C terminal domain, when compared to the overall length of the protein thread, was a sensation at the time and was published in the renowned scientific journal Nature.

Now the same group of researchers has put in place a further piece in the spider silk puzzle. They showed that the other end of the long thread, the so-called N terminal domain, plays an important role in the design of strong threads with great tensile strength. This time, the scientists investigated the head ends of the spider silk proteins of the "black widow" (Latrodectus hesperus). The result: The N terminal head ends exist in the silk duct as single strands (momomers). Only in the sinning passage are the head to tail pairs (dimers) formed.

The process of laying together is regulated via the change in pH values and salt concentrations between the silk duct and the spinning canal. In the silk duct, a neutral pH value of 7.2 and a high salt concentration prevent the N terminal head ends from combining. In the spinning passage, however, the environment becomes acidic (pH value around 6.2) the salt is removed. Now the ends can come together. In this process, the N terminal ends connect to the respective other ends – a practically endless chain of linked up spider silk proteins is formed. "In our work we were able to show, in addition to our previous research, that both the pH value and the salt concentration influence the monomer-dimer balance," says Franz Hagen, corresponding author of the study, in summing up the results. "Both factors influence the formation of dimers and thus the efficient cross-linking to very long silk proteins."

Ultimately, this cross-linking is what gives the spider silk threads their enormous tensile strength. The small crystallites first formed in the parallel cross-linking of the protein chains following the controlled unfolding of the C terminal domain are connected to each other via the N terminal domains of the spider silk protein to form a very long chain. "This is the effect that eventually explains the enormous tensile strength of the spider silk thread," says Kessler. To date, this ingenious form of cross-linking – called "multivalence" – has not been implemented in artificial polymers. "Most polymer chemists focus on the length of the thread. So far, no one has come up with the approach of cross-linking the ends of the threads and thereby opening the door to virtually unlimited lengths of polymer chains," beleives Kessler. These new findings may provide chemists with a model for manufacturing new materials with improved characteristics.

The scientists used the method of nuclear magnetic resonance (NMR) to analyze the structure of spider silk. Segments of spider silk are dissolved under conditions similar to those found in spider organs and exposed to radio wave impulses in a very strong magnetic field. The scientists can deduce the exact molecular structure from the "response" of the molecules. Using this method, environmental influences (e.g. salt concentration and pH value) can be studied accurately under simulated natural conditions. The development and application of NMR methods to biomolecules has been a longstanding focus of the Bavarian NMR Center in Garching.

More information: F. Hagn, C. Thamm, T. Scheibel, H. Kessler; pH Dependent Dimerisation and Salt Dependent Stabilisation of the N-terminal Domain of Spider Dragline Silk - Implications for Fibre Formation, Angew. Chem. Int. Ed. 2011, 50, 310-313.

Provided by Technische Universitaet Muenchen

Melanin's 'trick' for maintaining radioprotection studied

Sunbathers have long known that melanin in their skin cells provides protection from the damage caused by visible and ultraviolet light. More recent studies have shown that melanin, which is produced by multitudes of the planet's life forms, also gives some species protection from ionizing radiation. In certain microbes, in particular some organisms from near the former nuclear reactor facilities in Chernobyl, melanin has even been linked to increased growth in the presence of ionizing radiation.

Research at the U.S. Department of Energy's Savannah River National Laboratory, in collaboration with the Albert Einstein College of Medicine, has provided insights into the electrochemical mechanism that gives the complex polymer known as melanin its long-term radioprotective properties, with a goal of using that knowledge to develop materials that mimic those natural properties.

A recent article in the journal Bioelectrochemistry (Bioelectrochemistry 82 (2011) 69-73) relates how the researchers established that ionizing radiation interacts with melanin to alter its oxidation-reduction potential, resulting in electric current production.

Radiation causes damage by stripping away electrons from its target. "Over time, as melanin is bombarded with radiation and electrons are knocked away, you would expect to see the melanin become oxidized, or bleached out, and lose its ability to provide protection," said Dr. Charles Turick, Science Fellow with SRNL, "but that's not what we're seeing. Instead, the melanin continuously restores itself."

The team's research took them one step closer to understanding that self-restoration mechanism. They demonstrated that melanin can receive electrons, countering the oxidizing effects of the gamma radiation. The work showed, for the first time, that constant exposure of melanin to gamma radiation results in electric current production.

Mimicking that ability would be useful, for example, in the space industry, where satellites and other equipment are exposed to high levels of radiation for long spans of time. "Looking at materials, a constantly gamma radiation-oxidized electrode consisting in part of melanin would continuously accept electrons, thereby resulting in a current response," Turick said. "If we could understand how that works, we could keep that equipment working for a very long time."

Provided by Savannah River National Laboratory

Kinder, gentler cell capture method could aid medical research

 A research team at the National Institute of Standards and Technology (NIST) has come up with a potential solution to a two-pronged problem in medical research: How to capture cells on a particular spot on a surface using electric fields and keep them alive long enough to run experiments on them.

Their method, which involves innovations upon conventional cell-capture techniques, has already proved effective in creating arrays of human liver cells and mouse pluripotent cells—which, similar to stem cells, can develop into more than one cell type.

"The technique could prove valuable for learning about how cells communicate and differentiate," says NIST chemist Darwin Reyes. "We think this method could provide an effective way to selectively induce cells to differentiate and watch their behavior as they develop."

Adherent cells need to be attached to a surface to survive, and one common way of getting them there is by using a technique called dielectrophoresis (DEP), which Reyes says is not necessarily the best for cells' health. A batch of cells is placed into a fluid medium that has low electrical conductivity—sucrose in water, for example—and then subjected to an electric field that attracts the cells to a nearby surface. But the DEP process requires the cells to spend between 20 and 30 minutes in the medium, which appears to cause problems when the cells are trying to attach to the surface.

"Cells typically die rather soon after that much time exposed to the sucrose, since they cannot attach to the surface," Reyes says. "It's tough to run useful experiments if you only have a short window of opportunity."

The team experimented with different materials before finding that they could use a layer of substance called polyelectrolyte that has its own positive electric charge, which attracts the cells quickly. Before depositing this material, they laid down a thin layer of natural protein called fibronectin that helps cells to survive once they stick. With this new hybrid surface, the cells need spend only about four minutes in the fluid before they are returned to a more nurturing medium that helps them grow and attach better. As a result, the cells can survive on the surface for a week or more.

Because of their success in creating arrays of neural cells, the team has recently started to pattern liver cells as well. Combining liver cells with this technique could be useful in toxicology studies, Reyes suggests. "The liver is made up of several types of cells that work together," he says. "Creating arrays of them with certain cells positioned in particular locations could help us study how each of them might contribute to the overall process of filtering out a toxin from the bloodstream."

More information: D.R. Reyes, et al. Hybrid cell adhesive material for instant dielectrophoretic cell trapping and long-term cell function assessment. Langmuir, 2011, 27, 10027-10034, DOI: 10.1021/la200762j

Provided by National Institute of Standards and Technology (news : web)

Scientists uncover new factor in HIV infection

A George Mason University researcher team has revealed the specific process by which the HIV virus infects healthy T cells—a process previously unknown. The principal investigator, HIV researcher Yuntao Wu, says he hopes this breakthrough will start a new line on inquiry into how researchers can use this knowledge to create drugs that could limit or halt HIV infection.

Wu, a professor of molecular and microbiology at Mason, published these findings in an April 2011 edition of the Journal of Biological Chemistry, along with researchers Paul J. Vorster, Jia Guo, Alyson Yoder, Weifeng Wang, Yanfang Zheng, Dongyang Yu and Mark Spear from Mason's National Center for Biodefense and Infectious Diseases and the Department of Molecular and Microbiology and Xuehua Xu from Georgetown University School of Medicine's Department of Oncology.

This paper outlined a new understanding on how T cells—which are the target cells that the HIV virus infects—move and migrate when hijacked by the virus.

"The discovery adds to our understanding of how HIV initiates the infection of human T cells, which leads to their eventual destruction and the development of AIDS," Wu says.

Researchers and doctors have known for some time that the HIV virus, rather than directly killing healthy T cells, actually hijacks them. This eventually leads to their destruction. So the virus essentially turns the infected T cells (also known as CD4T cells or helper T cells) into a factory for creating even more HIV. Learning more about how the cells are infected could be a key step toward figuring out how to stop infection altogether.

Wu's latest discovery builds upon his previous work, published in the journal Cell in 2008, which described the basic process of how HIV infects T cells. After discovering that cofilin—a protein used to cut through a cell's outer layer, or cytoskeleton—is involved in HIV infection, Wu's new research provides the detailed framework for this process.

This new factor is called LIM domain kinase, or LIMK. The researchers discovered that LIMK triggers a cell to move, almost acting like a propeller. This cell movement is essential for HIV infection. This discovery marks the first time that a research team has uncovered the involvement of LIMK in HIV infection.

Building upon these results, the researchers then used a drug to trigger similar LIMK activation and found that it increased infection of T cells. Of course, the researchers ultimately want to decrease the infection of T cells—so they worked backwards and found something very promising.

"When we engineered the cell to inhibit LIMK activity, the cell became relatively resistant to HIV infection," says Wu. In other words, the researchers engineered human T cells that were not easily infected by HIV. This finding suggests that, in the future, drugs could be developed based on LIMK inhibition.

And while there are currently no medical drugs available to inhibit LIMK, Wu hopes this is a developing area in potential new therapeutic targets. One advantage of using this kind of therapy over the current medication available to those with HIV is that it's more difficult for the HIV virus to generate resistance to treatment, Wu explains.

Wu's team continues its work on decoding this complicated process, and he stresses that there is still much to be done.

"These findings are certainly exciting, and are an emerging research field that we are proud to have established three years ago with the publication of our Cell paper," he says. "We will continue to study the molecular details and to use those discoveries to develop new diagnostic and therapeutic tools to monitor and treat HIV-mediated CD4 T cell dysfunction and depletion."

Provided by George Mason University