New light on detection of bacterial infection: Polymers fluoresce in the presence of bacteria

 Researchers at the University of Sheffield have developed polymers that fluoresce in the presence of bacteria, paving the way for the rapid detection and assessment of wound infection using ultra-violet light.

When contained in a gel and applied to a wound, the level of fluorescence detected will alert clinicians to the severity of infection. The polymers are irreversibly attached to fragments of antibiotics, which bind to either gram negative or gram positive bacteria -- both of which cause very serious infections -- informing clinicians as to whether to use antibiotics or not, and the most appropriate type of antibiotic treatment to prescribe. The team also found that they could use the same gels to remove the bacteria from infected wounds in tissue engineered human skin.

Professor Sheila MacNeil, an expert in tissue engineering and wound healing, explained: "The polymers incorporate a fluorescent dye and are engineered to recognise and attach to bacteria, collapsing around them as they do so. This change in polymer shape generates a fluorescent signal that we´ve been able to detect using a hand-held UV lamp."

"The availability of these gels would help clinicians and wound care nurses to make rapid, informed decisions about wound management, and help reduce the overuse of antibiotics," added project lead Dr Steve Rimmer.

Currently, determining significant levels of bacterial infection involves swabbing the wound and culturing the swabs in a specialist bacteriology laboratory with results taking several days to be available. The team is confident that its technology can ultimately reduce the detection of bacterial infection to within a few hours, or even less.

The research has already demonstrated that the polymer (PNIPAM), modified with an antibiotic (vancomycin) and containing a fluorescent dye (ethidium bromide), shows a clear fluorescent signal when it encounters gram negative bacteria. Other polymers have been shown to respond to S. aureus, a gram positive bacteria. These advances mean that a hand-held sensor device can now be developed to be used in a clinical setting.

The research is the result of a three-year project which started in 2006, part-funded by the Engineering and Physical Sciences Research Council (EPSRC) and the Defence Science and Technology Laboratory (Dstl) -- an agency of the Ministry of Defence, interested in the medical application of the research in battlefield conditions, and a subsequent EPSRC funded PhD studentship.

The team is also investigating whether using a sophisticated technique called fluorescence Non Radiative Energy Transfer (NRET) to generate the light signal could enable a highly refined sensor technology that could have applications in other areas.

"For example, we think that NRET could be very useful in an anti-terrorist and public health capacity, detecting pathogen release or bacterial contamination, whether accidental or deliberate," says Dr Rimmer. "NRET also allows us to learn more about how the polymers collapse around the bacteria, which is important in developing our understanding of how bacteria interact with these novel responsive polymers."

The team is interested in talking to potential partners to take this technology forward.

Story Source:

The above story is reprinted (with editorial adaptations ) from materials provided by University of Sheffield.

'Inexhaustible' source of hydrogen may be unlocked by salt water, engineers say

 A grain of salt or two may be all that microbial electrolysis cells need to produce hydrogen from wastewater or organic byproducts, without adding carbon dioxide to the atmosphere or using grid electricity, according to Penn State engineers.

"This system could produce hydrogen anyplace that there is wastewater near sea water," said Bruce E. Logan, Kappe Professor of Environmental Engineering. "It uses no grid electricity and is completely carbon neutral. It is an inexhaustible source of energy."

Microbial electrolysis cells that produce hydrogen are the basis of this recent work, but previously, to produce hydrogen, the fuel cells required some electrical input. Now, Logan, working with postdoctoral fellow Younggy Kim is using the difference between river water and seawater to add the extra energy needed to produce hydrogen.

Their results, published Sept. 19 in the Proceedings of the National Academy of Sciences, "show that pure hydrogen gas can efficiently be produced from virtually limitless supplies of seawater and river water and biodegradable organic matter."

Logan's cells were between 58 and 64 percent efficient and produced between 0.8 to 1.6 cubic meters of hydrogen for every cubic meter of liquid through the cell each day. The researchers estimated that only about 1 percent of the energy produced in the cell was needed to pump water through the system.

The key to these microbial electrolysis cells is reverse-electrodialysis or RED that extracts energy from the ionic differences between salt water and fresh water. A RED stack consists of alternating ion exchange membranes -- positive and negative -- with each RED contributing additively to the electrical output.

"People have proposed making electricity out of RED stacks," said Logan. "But you need so many membrane pairs and are trying to drive an unfavorable reaction."

For RED technology to hydrolyze water -- split it into hydrogen and oxygen -- requires 1.8 volts, which would in practice require about 25 pairs of membrane sand increase pumping resistance. However, combining RED technology with exoelectrogenic bacteria -- bacteria that consume organic material and produce an electric current -- reduced the number of RED stacks to five membrane pairs.

Previous work with microbial electrolysis cells showed that they could, by themselves, produce about 0.3 volts of electricity, but not the 0.414 volts needed to generate hydrogen in these fuel cells. Adding less than 0.2 volts of outside electricity released the hydrogen. Now, by incorporating 11 membranes -- five membrane pairs that produce about 0.5 volts -- the cells produce hydrogen.

"The added voltage that we need is a lot less than the 1.8 volts necessary to hydrolyze water," said Logan. "Biodegradable liquids and cellulose waste are abundant and with no energy in and hydrogen out we can get rid of wastewater and by-products. This could be an inexhaustible source of energy."

Logan and Kim's research used platinum as a catalyst on the cathode, but subsequent experimentation showed that a non-precious metal catalyst, molybdenum sulfide, had a 51 percent energy efficiency. The King Abdullah University of Science and Technology supported this work.

Story Source:

The above story is reprinted (with editorial adaptations ) from materials provided by Penn State.

Journal Reference:

Younggy Kim, Bruce E. Logan. Hydrogen production from inexhaustible supplies of fresh and salt water using microbial reverse-electrodialysis electrolysis cells. Proceedings of the National Academy of Sciences, 2011; DOI: 10.1073/pnas.1106335108

Scientists solve long-standing plant biochemistry mystery

 Scientists at the U.S. Department of Energy's (DOE) Brookhaven National Laboratory and collaborators at the Karolinska Institute in Sweden have discovered how an enzyme "knows" where to insert a double bond when desaturating plant fatty acids. Understanding the mechanism -- which relies on a single amino acid far from the enzyme's active site -- solves a 40-year mystery of how these enzymes exert such location-specific control.

The work, published in the Proceedings of the National Academy of Sciences the week of September 19, 2011, may lead to new ways to engineer plant oils as a renewable replacement for petrochemicals.

"Plant fatty acids are an approximately $150-billion-dollar-a-year market," said Brookhaven biochemist John Shanklin, lead author on the paper. "Their properties, and therefore their potential uses and values, are determined by the position of double bonds in the hydrocarbon chains that make up their backbones. Thus the ability to control double bond positions would enable us to make new designer fatty acids that would be useful as industrial raw materials."

The enzymes responsible for double-bond placement, called desaturases, remove hydrogen atoms and insert double bonds between adjacent carbon atoms at specific locations on the hydrocarbon chains. But how one enzyme knows to insert the double bond at one location while a different but closely related enzyme inserts a double bond at a different site has been a mystery.

"Most enzymes recognize features in the molecules they act on that are very close to the site where the enzyme's action takes place. But all the carbon-hydrogen groups that make up fatty-acid backbones are very similar with no distinguishing features -- it's like a greasy rope with nothing to hold onto," said Shanklin.

In describing his group's long-standing quest to solve the desaturation puzzle, Shanklin quotes Nobel laureate Konrad Bloch, who observed more than 40 years ago that such site-specific removal of hydrogen "would seem to approach the limits of the discriminatory power of enzymes."

Shanklin and his collaborators approached the problem by studying two genetically similar desaturases that act at different locations: a castor desaturase that inserts a double bond between carbon atoms 9 and 10 in the chain (a 'delta-9' desaturase); and an ivy desaturase that inserts a double bond between carbon atoms 4 and 5 (delta-4). They reasoned that any differences would be easy to spot in such extreme examples.

But early attempts to find a telltale explanation -- which included detailed analyses of the two enzymes' atomic-level crystal structures -- turned up few clues. "The crystal structures are almost identical," Shanklin said.

The next step was to look at how the two enzymes bind to their substrates -- fatty acid chains attached to a small carrier protein. First the scientists analyzed the crystal structure of the castor desaturase bound to the substrate. Then they used computer modeling to further explore how the carrier protein "docked" with the enzyme.

"Results of the computational docking model exactly matched that of the real crystal structure, which allows carbon atoms 9 and 10 to be positioned right at the enzyme's active site," Shanklin said.

Next the scientists modeled how the carrier protein docked with the ivy desaturase. This time it docked in a different orientation that positioned carbon atoms 4 and 5 at the desaturation active site. "So the docking model predicted a different orientation that exactly accounted for the specificity," Shanklin said.

To identify exactly what was responsible for the difference in binding, the scientists then looked at the amino acid sequence -- the series of 360 building blocks that makes up each enzyme. They identified amino acid locations that differ between delta-9 and delta-4 desaturases, and focused on those locations that would be able to interact with the substrate, based on their positions in the structural models.

The scientists identified one position, far from the active site, where the computer model indicated that switching a single amino acid would change the orientation of the bound fatty acid with respect to the active site. Could this distant amino-acid location remotely control the site of double bond placement?

To test this hypothesis, the scientists engineered a new desaturase, swapping out the aspartic acid normally found at that location in the delta-9 castor desaturase for the lysine found in the delta-4 ivy desaturase. The result: an enzyme that was castor-like in every way, except that it now seemed able to desaturate the fatty acid at the delta-4 carbon location. "It's quite remarkable to see that changing just one amino acid could have such a striking effect," Shanklin said.

The computational modeling helped explain why: It showed that the negatively charged aspartic acid in the castor desaturase ordinarily repels a negatively charged region on the carrier protein, which leads to a binding orientation that favors delta-9 desaturation; substitution with positively charged lysine results in attraction between the desaturase and carrier protein, leading to an orientation that favors delta-4 desaturation.

Understanding this mechanism led Ed Whittle, a research associate in Shanklin's lab, to add a second positive charge to the castor desaturase in an attempt to further strengthen the attraction. The result was a nearly complete switch in the castor enzyme from delta-9 to delta-4 desaturation, adding compelling support for the remote control hypothesis.

"I really admire Ed's persistence and insight in taking what was already a striking result and pushing it even further to completely change the way this enzyme functions," Shanklin said.

"It's very rewarding to have finally solved this mystery, which would not have been possible without a team effort drawing on our diverse expertise in biochemistry, genetics, computational modeling, and x-ray crystallography.

"Using what we've now learned, I am optimistic we can redesign enzymes to achieve new desirable specificities to produce novel fatty acids in plants. These novel fatty acids would be a renewable resource to replace raw materials now derived from petroleum for making industrial products like plastics," Shanklin said.

This work was funded by the DOE Office of Science. Additional collaborators include: Jodie Guy, Martin Moche, and Ylva Lindqvist of the Karolinska Institute, and Johan Lengqvist, now at AstraZeneca R&D in Sweden. The scientists analyzed crystal structures at several synchrotrons including: the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory, the European Synchrotron Radiation Facility (ESRF) in France, the Dutch Electron Synchrotron (DESY), and the MAX-lab National Laboratory for Synchrotron Radiation in Sweden.

Story Source:

The above story is reprinted (with editorial adaptations) from materials provided by DOE/Brookhaven National Laboratory.

Journal Reference:

Jodie E. Guy, Edward Whittle, Martin Moche, Johan Lengqvist, Ylva Lindqvist, John Shanklin. Remote control of regioselectivity in acyl-acyl carrier protein-desaturases. Proceedings of the National Academy of Sciences, 2011; DOI: 10.1073/pnas.1110221108

Researchers power line-voltage light bulb with nanotube wire

 Cables made of carbon nanotubes are inching toward electrical conductivities seen in metal wires, and that may light up interest among a range of industries, according to Rice University researchers.

A Rice lab made such a cable from double-walled carbon nanotubes and powered a fluorescent light bulb at standard line voltage -- a true test of the novel material's ability to stake a claim in energy systems of the future.

The work appears this week in the Nature journal Scientific Reports.

Highly conductive nanotube-based cables could be just as efficient as traditional metals at a sixth of the weight, said Enrique Barrera, a Rice professor of mechanical engineering and materials science. They may find wide use first in applications where weight is a critical factor, such as airplanes and automobiles, and in the future could even replace traditional wiring in homes.

The cables developed in the study are spun from pristine nanotubes and can be tied together without losing their conductivity. To increase conductivity of the cables, the team doped them with iodine and the cables remained stable. The conductivity-to-weight ratio (called specific conductivity) beats metals, including copper and silver, and is second only to the metal with highest specific conductivity, sodium.

Yao Zhao, who recently defended his dissertation toward his doctorate at Rice, is the new paper's lead author. He built the demo rig that let him toggle power through the nanocable and replace conventional copper wire in the light-bulb circuit.

Zhao left the bulb burning for days on end, with no sign of degradation in the nanotube cable. He's also reasonably sure the cable is mechanically robust; tests showed the nanocable to be just as strong and tough as metals it would replace, and it worked in a wide range of temperatures. Zhao also found that tying two pieces of the cable together did not hinder their ability to conduct electricity.

The few centimeters of cable demonstrated in the present study seems short, but spinning billions of nanotubes (supplied by research partner Tsinghua University) into a cable at all is quite a feat, Barrera said. The chemical processes used to grow and then align nanotubes will ultimately be part of a larger process that begins with raw materials and ends with a steady stream of nanocable, he said. The next stage would be to make longer, thicker cables that carry higher current while keeping the wire lightweight. "We really want to go better than what copper or other metals can offer overall," he said.

The paper's co-authors are Tsinghua researcher Jinquan Wei, who spent a year at Rice partly supported by the Armchair Quantum Wire Project of Rice University's Smalley Institute for Nanoscale Science and Technology; Robert Vajtai, a Rice faculty fellow in mechanical engineering and materials science; and Pulickel Ajayan, the Benjamin M. and Mary Greenwood Anderson Professor of Mechanical Engineering and Materials Science and professor of chemistry and chemical and biomolecular engineering.

The Research Partnership to Secure Energy for America, the Department of Energy and Air Force Research Laboratory supported the project.

Story Source:

The above story is reprinted (with editorial adaptations ) from materials provided by Rice University.

Journal Reference:

Yao Zhao, Jinquan Wei, Robert Vajtai, Pulickel M. Ajayan, Enrique V. Barrera. Iodine doped carbon nanotube cables exceeding specific electrical conductivity of metals. Scientific Reports, 2011; 1 DOI: 10.1038/srep00083