Chemists devise a way to create a five point knotted molecule

Called a pentafoil, the five point knot is the most complex kind of molecule synthesized from other building blocks, other than those found in DNA, and having a means for building them could lead to all sorts of new materials that could be both strong and flexible.

To build the molecule, the team started with a negatively charged chloride ion, to serve as a pulling force, or anchor. They then added other parts, such as iron ions with a positive charge, and chains of carbon atoms. They then chemically “programmed” the whole works to assemble itself into the pentafoil, with five chains looped over and under one another and connected to form one single knotted strand, with a single chloride ion sitting squarely in the center holding the whole knot together. The finished product is made up of just 160 atoms and very much resembles a traditional two-dimensional five pointed star.

As an interesting side note, the researchers found that if they removed the single chloride ion after the knot was completed, they were left with a molecule that was hungry for that missing ion, which could mean they’ve found a new type of chlorine sensor.

In devising a means to create a pentafoil, the researchers have created not just a new type of man-made molecule, but a blueprint for creating other types of knotted molecules which could lead to all sorts of new and exotic materials.

More information: A synthetic molecular pentafoil knot, Nature Chemistry 4, 15–20 (2012) doi:10.1038/nchem.1193

Abstract
Knots are being discovered with increasing frequency in both biological and synthetic macromolecules and have been fundamental topological targets for chemical synthesis for the past two decades. Here, we report on the synthesis of the most complex non-DNA molecular knot prepared to date: the self-assembly of five bis-aldehyde and five bis-amine building blocks about five metal cations and one chloride anion to form a 160-atom-loop molecular pentafoil knot (five crossing points). The structure and topology of the knot is established by NMR spectroscopy, mass spectrometry and X-ray crystallography, revealing a symmetrical closed-loop double helicate with the chloride anion held at the centre of the pentafoil knot by ten CH···Cl– hydrogen bonds. The one-pot self-assembly reaction features an exceptional number of different design elements—some well precedented and others less well known within the context of directing the formation of (supra)molecular species. We anticipate that the strategies and tactics used here can be applied to the rational synthesis of other higher-order interlocked molecular architectures.

? 2011 PhysOrg.com

Biochemists develop promising new treatment direction for rare metabolic diseases

People born with Fabry disease have a faulty copy of a single gene that codes for the alpha-galactosidase (?-GAL) enzyme, one of the cell's "recycling" machines. When it performs normally, ?-GAL breaks down an oily lipid known as GB3 in the cell's recycling center, or lysosome. But when it underperforms or fails, Fabry symptoms result. Patients may survive to adulthood, but the disorder leads to toxic lipid build-up in blood vessels and organs that compromise kidney function or lead to heart disease, for example.

The faulty gene causes its damage by producing a misfolded protein, yielding an unstable, poorly functioning ?-GAL enzyme. Like origami papers, these proteins are unfolded to start and only become active when folded into precise shapes. At present, enzyme replacement therapy (ERT) is the only FDA-approved treatment for such lysosomal storage disorders as Fabry, Pompe and Gaucher diseases, but ERT requires a complicated and expensive process to purify and replace the damaged ?-GAL enzyme, and it must be administered by a physician.

Instead of replacing the damaged enzyme, an alternative route called pharmacological chaperone (PC) therapy is currently in Phase III clinical trials for Fabry disease. It relies on using smaller, "chaperone" molecules to keep proteins on the right track toward proper folding, but their biochemical mechanism is not well understood, says Garman.

Now, he and colleagues report results of a thorough exploration at the atomic level of the biochemical and biophysical basis of two small molecules for potentially stabilizing the ?-GAL enzyme. He says their use in PC therapy could one day be far less expensive than the current standard, ERT, and can be taken orally.

This work, which improves knowledge of a whole class of molecular chaperones, represents the centerpiece of UMass Amherst student Abigail Guce's doctoral thesis and was supported by the National Institutes of Health. Other members of the team are graduate students Nat Clark and Jerome Rogich.

"The interactions we looked at are exactly the things occurring in the clinical trial right now," Garman says. Further, "the same concept is now being applied to other protein-folding diseases such as Parkinson's and Alzheimer's disease. Many medical researchers are trying to keep proteins from misfolding by using small chaperone molecules. Our studies have definitely advanced the understanding of how to do that."

In their current paper, Garman and colleagues compare the ability of two small chaperone molecules, galactose and 1-deoxygalactononjirimycin (DGJ) to stabilize the ?-GAL protein, to help it resist unfolding in different conditions such as high temperature and different pH levels.

They found that each chaperone has very different affinities: DGJ binds tightly and galactose binds loosely to the ?-GAL, yet they differ in only two atomic positions. "Tight is better, because you can use less drug for treatment," Garman says. "We now can explain DGJ's high potency, its tight binding, down to individual atoms."

In earlier studies as in the current work, the UMass Amherst team used their special expertise in X-ray crystallography to create three-dimensional images of all atoms in the protein to understand how it carries out its metabolic mission. They also found a new binding site for small molecules on human ?-GAL that had never been observed before.

Crystallography on the two chaperones bound to the ?-GAL enzyme showed that a single interaction between the enzyme and DGJ was responsible for DGJ's high affinity for the enzyme. Other experiments also showed the ability of the 11- and 12-atom chaperones to protect the large, 6,600-atom ?-GAL from unfolding and degradation.

For the first time, by making a single change in one amino acid in protein, they forced the DGJ to bind weakly, indicating that one atomic interaction is responsible for DGJ's high affinity.

"It was surprising to find these two small molecules that look very much the same have very different affinities for this enzyme," says Garman, "and we now understand why. The iminosugar DGJ has high potency due to a single ionic interaction with ?-GAL. Overall, our studies show that this small molecule keeps the enzyme from unfolding, or when it unfolds, the process happens more slowly, all of which you need in treating disease."

Provided by University of Massachusetts at Amherst

No more free rides for 'piggy-backing' viruses

The findings open the door to the development of new drugs to combat these deadly viruses that infect more than 180 million people worldwide.

The team of international scientists led by and Professor Gideon Davies from the University of York and Associate Professor Spencer Williams from the University of Melbourne, studied bacterial endomannosidase as a model for the same human enzyme and successfully determined the three dimensional structure of the enzyme using state of the art synchrotron technology.

Professor Davies, of the Department of Chemistry at York, said that knowing the structure of the enzyme revealed details on how viruses play biological "piggy-back", borrowing our cellular machinery to replicate and cause disease

"If we understand how the viruses use our enzymes, we can develop inhibitors that block the pathway they require, opening the door to drug developments," he said.

In the past the problem has been that this group of viruses including HIV, Hepatitis C, Dengue Fever and West Nile virus, are able to bypass the main pathway if inhibited and replicate via a second pathway using this enzyme. Thus for a treatment to be effective, both pathways need to be blocked.

"It was already known how to block the main pathway for these viruses but until now, this endomannosidase bypass pathway has proved a considerable challenge to study," Professor Davies said.

Dr Williams said: "Combining international resources and expertise, we were able to determine the endomannosidase structure and this has revealed how we can block the bypass route, stopping the viruses from hijacking human enzymes."

Professor Davies added: "We hope that the work will lead beyond viruses and will point the way towards similar treatments for other diseases including cancer."

The study is published in the Proceedings of the National Academy of Sciences (PNAS) this week.

More information: ‘Structural and mechanistic insight into N-glycan processing by endo-?-mannosidase’ http://www.pnas.or … s.1111482109

Provided by University of York

Chemistry trick renews hope against killer diseases

Now a Danish chemist has pioneered a novel way to battle multidrug resistant bacteria. By tweaking a well known psychoactive drug he revitalizes worn-out drugs like sulpha and penicillin.

Chemist Jorn Bolstad Christensen of the University of Copenhagen has just patented the use of schizophrenia medication Thioridazin in boosting the effect of antibiotics. Christensen is an associate professor at the Department of Chemistry, University of Copenhagen, and in the lab he started investigating how the schizophrenia drug might bother bacteria but not humans.

"Thioridazin blocks the capacity of bacteria to cleanse themselves of antibiotics. We knew that before starting. But I wanted to remove the action of the drug in the brain so that mortally ill tuberculosis patients wouldn't have to contend with psychoactive effects as a part of their cure," explains Christensen, who none the less had to come to terms with a much bigger threat as well.

Bacteria such as those responsible for tuberculosis, staphylococcus and enterococcus get rid of antibiotics using their so called efflux-pump. A mechanism which simply pumps the active substance out of the cell before it has an opportunity to do harm. A substance which blocks the pump should ensure that any antibiotic stays inside the bacteria long enough to kill it. There's just one tiny problem. Human cells have efflux pumps as well. And we wouldn't want these blocked.

"The task was to find a substance that will kill bacteria, but not the patients taking the cure. Thioridazin was a good candidate because it's been in use for decades. We could be pretty certain that it wouldn't have any serious side effects," says professor Christensen, who predicts testing the new drug in humans within just a year, as it's already been approved for other medical uses.

Though the risk of blocking the efflux pump of human cells appeared minimal Professor Christensen still needed to minimize the psychoactive effects of the drug. This was where his chemical expertise became indispensable. Chemically Thioridazin consists of two half molecules which are perfect mirror images. One of these mirror- or isomeric forms affects the brain less than the other, so the question was whether bacteria would know the difference.

Doctors and researchers Jette Kristiansen and Oliver Hendricks of Southern University Denmark who are co-holders of the patent have conducted microbiological trials proving that the efflux pump of bacteria stayed blocked regardless of which isomer was used. These results open up a brand new method for combating problems of multidrug resistance.

Provided by University of Copenhagen

NSF turns to ancient pottery to improve modern heat resistant ceramics

In order for space vehicles to work properly, they have to be able to withstand the cold of space, which can be as low as 250 degrees Fahrenheit below zero. Then, some vehicles have to be able to withstand the extreme heat of reentry, which can be as hot as 3000 degrees. Hot enough to melt most any metal. As most know, the Space Shuttle was fitted with ceramic tiles on its underbelly to keep the vehicle from overheating as it came back to Earth, which was made all the more apparent when damage to the tiles resulted in the loss of Columbia in 2003. But ceramics are used in other components as well, and will be needed as more ambitious projects are undertaken in the future. Equally important is the ability of ceramics to remain chemically unchanged when subjected to such temperature variations. Such properties allow for the construction of components that minimize expansion and contraction under such stresses, which can be critical for long term operations in space. This is why the NSF has turned to research scientists to see if the ancient Greeks can provide some insight.

Such research will involve using something called x-ray absorption near edge structure (XANES) - which is a special type of spectroscopy, along with other types of x-ray techniques, to find out what has gone on with iron oxidation in the pottery under study. What’s needed is a better understanding of the molecular structure of iron minerals that were used to make the pottery to help researchers in designing newer and better types of ceramics for future space missions, whether manned or otherwise.

What’s interesting is that it is apparently the degree to which the iron in the ancient ceramic pottery oxidized that caused the distinctive red and black colorations that made it so attractive to those that worked with it all those years ago.