Scientists probe form, function of mysterious protein

Using a combination of laboratory experiments and computer modeling, scientists from Rice University and the University of California, San Diego (UCSD) have deciphered part of mitoNEET's movements to get a better understanding of how it handles its potentially toxic payload of iron and sulfur. Their research is described this week in the Proceedings of the National Academy of Sciences.

"We scrutinize proteins with an unconventional approach," said José Onuchic, Rice's Harry C. and Olga K. Wiess Professor of Physics and Astronomy and co-director of the Center for Theoretical Biological Physics. "We use biophysics to probe biology rather than the other way around. Using computational theory, we find structures that are possible -- regardless of whether they've already been observed experimentally -- and we ask ourselves whether these structures might be biologically significant."

Study co-leader Patricia Jennings, professor of chemistry and biochemistry at UCSD, who has collaborated with Onuchic for 15 years, said they save a great deal of time by using structural biophysics to guide their experiments on a wide variety of targets. For example, Jennings' laboratory determined less than five years ago that mitoNEET contained a novel folded structure. Since then, her lab has been using insights gained from static and dynamic snapshots of the protein to guide biological and biochemical studies.

"I think people forget that proteins are machines with moving parts," said study lead author Elizabeth Baxter, a UCSD graduate student who works under the guidance of both Onuchic and Jennings. "We start with the static snapshot and model in the functional motions."

MitoNEET, which binds to the diabetes drug, Actos, immediately caught the attention of researchers when it was discovered. It has a unique ability to bind and store iron-based molecules in an iron-sulfur cluster. Iron is an essential element for all life, but it is also highly toxic, and mitoNEET is the only iron-handling protein that is known to sit on the wall of the mitochondria, one of the key structures inside a cell.

The protein's biological functions are still being unraveled. Interestingly, scientists have shown that mitoNEET sits on the outer mitochondrial wall with its potentially toxic payload of iron-sulfur molecules facing toward the cell's cytoplasm, the gel-like fluid that fills the cell. Discovery of the unique binding mode of the protein's iron-sulfur cluster led the Jennings group to show that the cluster can be delivered into the mitochondria. In addition, its sister protein interacts with proteins that participate in apoptosis -- the process cells use to kill themselves when they are no longer viable.

"I think mitoNEET is a protein that could be your best friend or your worst enemy," Jennings said. "There's some evidence that it may act as a sensor for oxidative stress and that it can lose its toxic iron-sulfur cluster under stress conditions. Depending upon where the iron ends up, that could lead to drastic problems inside the cell."

Proteins are strands of amino acids that are produced from DNA blueprints, but their shapes can provide important clues about their function. To find out how mitoNEET's control and release of its iron-sulfur payload might be related to its shape, Baxter used computer simulations to study how the protein folds, as well as the functional motions of two similar shapes that could be biologically important. In one of these shapes, there is a slight intertwining of two arms that extend away from the iron-cluster pocket. In the other, the arms also extend but are not intertwined.

Baxter found that both conformations were physically possible. She also found the protein could switch between the "strand-swapped" and "strand-unswapped" conformations without entirely unfolding. Moreover, this change in the twining of the arms was shown to alter the shape of the critical pocket that holds the iron-sulfur cluster; this makes the cluster more likely to be inserted or released in situations where the arms are untwined.

Like the magician using misdirection, the loosening of the grip on the cluster is subtle and happens in a different location than the flurry of arm motions. Jennings said it's the kind of thing that could easily be missed if the focus of the study were the cluster itself.

Onuchic said, "One of the advantages to our approach is that it allows us to look for relevant biophysical properties that control distant functional regions -- like mitoNEET's strand-swapping -- that can easily be missed with a more conventional approach."

More information: http://www.pnas.or … 109.abstract

Provided by Rice University (news : web)

Studying the chemistry as it happens in catalytic reactions

"Scientists have been trying for a long time to get something closer to a realistic environment with NMR data. This is the newest approach to doing that," said Dr. Charles Peden, a catalysis researcher in the Institute for Integrated Catalysis at PNNL who worked on the study.

From refining gasoline to manufacturing margarine, catalysts are involved in ~90% of all commercially produced chemical products. Making existing catalysts more effective or devising new ones could reduce costly inefficiencies in current processes, and could enable new commercial processes to produce fuels and chemicals. To improve existing and invent new catalysts and catalytic processes, scientists need data about the steps that occur during the reaction. With this new probe, scientists get that type of detailed information via NMR spectroscopy.

"NMR is a powerful technique.  Being able to apply it to catalytic reactions while they are occurring has been a really tough problem. This new in situ NMR probe lets us perform experiments we couldn't do before," said Dr. Jian Zhi Hu, the lead PNNL scientist on this study.

In a reaction that turns chemical A into chemical Z, many intermediates can be formed. Some of these can become undesirable waste products, but others eventually form the desired product.

The challenge with using NMR for catalysis centers on a technique called magic angle spinning. This method requires spinning the catalyst sample inside the NMR instrument. The scientists want the sample to represent how it exists during the actual industrial catalytic process. They want the reaction of gaseous feedstocks over the solid catalyst to be taking place right when they collect the NMR data.

"You need to spin the sample—often several thousand times a second, but it's not possible with gas lines directly attached," said Hu.

But, they really wanted the data.

So, working together in EMSL, they devised a new method that allowed them to use magic angle spinning on solid catalyst materials with the desired continuous flow of gaseous materials. They built an NMR probe, containing the mechanisms for magic angle sample spinning and an NMR transmitter and receiver, which fit into the heart of the spectrometer's superconducting magnet.  At one end of the probe is a heatable sample cell filled with a solid catalyst. This cell includes some special connections to gas lines that allow for introduction of fresh reactants into the sample chamber. In the chamber, the gas reacts with the catalyst. The gases, now containing varying concentrations of reactants and products, are pulled to a vacuum pump through the tube. The connections of the magic angle spinning cell to the gas lines are made in such a way as to minimize leakage while still allowing for high speed spinning.  Another important feature is that the probe allows for the use of a large catalyst sample volume for enhanced sensitivity.

"EMSL has a really fantastic technical team that can construct these probes," said Peden, who has conducted experiments at EMSL since the facility opened in 1997.

Next, the team used the device to follow several different catalytic reactions. For example, they followed a dehydration reaction that removes a water molecule from a 4-carbon alcohol. The team identified the type and number of different isomers of the product that formed during the catalytic reaction.

This new probe will soon be available to scientists through EMSL's user proposal process.

The scientists are conducting experiments on different catalysts and catalyzed reactions, working in collaboration with colleagues from the University of California at Berkeley. In addition, they are considering how to refine the probe, creating a next-generation device that can even more closely match industrial catalysis conditions.

More information: JZ Hu, et al. 2012. "A Large Sample Volume Magic Angle Spinning Nuclear Magnetic Resonance Probe for In Situ Investigations with Constant Flow of Reactants." Physical Chemistry Chemical Physics 14, 2137-2143. DOI: 10.1039/c1cp22692d

Provided by Pacific Northwest National Laboratory (news : web)

Cosmology in a Petri dish

To understand long-range interactions between particles at the micrometric scale, researchers utilize methods which are used to study the formation of our universe.

Scientists have found that micron-size particles which are trapped at fluid interfaces exhibit a collective dynamic that is subject to seemingly unrelated governing laws. These laws show a smooth transitioning from long-ranged cosmological-style gravitational attraction down to short-range attractive and repulsive forces. The study is by Johannes Bleibel from the Max Planck Institute for Intelligent Systems in Stuttgart, Germany, and his colleagues.

The authors used so-called colloidal particles that are larger than molecules but too small to be observed with the naked eye. These particles are adsorbed at the interface between two fluids and assembled into a monolayer. This constitutes a 2D model in which particles that are larger than a micron deform the interface through their own weight and generate an effective long-range attraction which looks like gravitation in 2D. Thus, the particles assemble in clusters.

To model long-range forces between particles, the researchers used numerical simulations based on random movement of particles, known as Brownian dynamics. Here, they took advantage of the formal analogy between so-called capillary attraction -- the long-ranged interaction through interface deformation -- and gravitational attraction. They used a particle-mesh method as employed in simulations of what are known as self-gravitating fluids, corresponding to the collapse of a system under its own gravity, traditionally used in cosmological studies.

The authors also found that this long-range interaction no longer matters beyond a certain length determined by the properties of both the particles and the interface, and short-range forces come into play. This means that for systems exceeding this length, particles first tend to self-assemble into several clusters which eventually merge into a single, large cluster.

The study of monolayer aggregates of micron-size colloids is used as a template for nanoparticles deposited onto substrates in nanotechnology applications.

Story Source:

The above story is reprinted from materials provided by Springer, via AlphaGalileo.

Note: Materials may be edited for content and length. For further information, please contact the source cited above.

Journal Reference:

Bleibel J, Dominguez A, Oettel M, Dietrich S. Collective dynamics of colloids at fluid interfaces. European Physical Journal E, 34:125 DOI: 10.1140/epje/i2011-11125-2

Oxygen molecule survives to enormously high pressures

Using computer simulations, a RUB researcher has shown that the oxygen molecule (O2) is stable up to pressures of 1.9 terapascal, which is about nineteen million times higher than atmosphere pressure. Above that, it polymerizes, i.e. builds larger molecules or structures.

"This is very surprising" says Dr. Jian Sun from the Department of Theoretical Chemistry. "Other simple molecules like nitrogen or hydrogen do not survive such high pressures." In cooperation with colleagues from University College London, the University of Cambridge, and the National Research Council of Canada, the researcher also reports that the behaviour of oxygen with increasing pressure is very complicated. It's electrical conductivity first increases, then decreases, and finally increases again. The results are published in Physical Review Letters.

Weaker bonds, greater stability

The oxygen atoms in the O2 molecule are held together by a double covalent bond. Nitrogen (N2), on the other hand, possesses a triple bond. "You would think that the weaker double bond is easier to break than the triple bond and that oxygen would therefore polymerize at lower pressures than nitrogen" says Sun. "We found the opposite, which is astonishing at first sight."

Coming together when pressure increases

However, in the condensed phase when pressure increases, the molecules become closer to each other. The research team suggests that, under these conditions, the electron lone pairs on different molecules repel one another strongly, thus hindering the molecules from approaching each other. Since oxygen has more lone pairs than nitrogen, the repulsive force between these molecules is stronger, which makes polymerization more difficult. However, the number of lone pairs cannot be the only determinant of the polymerization pressure. "We believe that it is a combination of the number of lone pairs and the strength of the bonds between the atoms," says Sun.

The many structures of oxygen

At high pressures, gaseous molecules such as hydrogen, carbon monoxide, or nitrogen polymerize into chains, layers, or framework structures. At the same time they usually change from insulators to metals, i.e. they become more conductive with increasing pressure. The research team, however, showed that things are more complicated with oxygen. Under standard conditions, the molecule has insulating properties. If the pressure increases, oxygen metallises and becomes a superconductor. With further pressure increase, its structure changes into a polymer and it becomes semi-conducting. If the pressure rises even more, oxygen once more assumes metallic properties, meaning that the conductivity goes up again. The metallic polymer structure finally changes into a metallic layered structure.

Inside planets

"The polymerization of small molecules under high pressure has attracted much attention because it helps to understand the fundamental physics and chemistry of geological and planetary processes" explains Sun. "For instance, the pressure at the centre of Jupiter is estimated to be about seven terapascal. It was also found that polymerized molecules, like N2 and CO2, have intriguing properties, such as high energy densities and super-hardness." Dr. Jian Sun joined the RUB-research group of Prof. Dr. Dominik Marx as a Humboldt Research Fellow in 2008 to work on vibrational spectroscopy of aqueous solutions. In parallel to this joint work in Solvation Science he developed independent research interests into high pressure chemical physics as an Early Career Researcher.

Story Source:

The above story is reprinted from materials provided by Ruhr-Universitaet-Bochum, via AlphaGalileo.

Note: Materials may be edited for content and length. For further information, please contact the source cited above.

Journal Reference:

Jian Sun, Miguel Martinez-Canales, Dennis Klug, Chris Pickard, Richard Needs. Persistence and Eventual Demise of Oxygen Molecules at Terapascal Pressures. Physical Review Letters, 2012; 108 (4) DOI: 10.1103/PhysRevLett.108.045503