SynBioFromLeukipposInstitute

The team of researchers at Saarland University, led by Professor of Condensed Matter Physics Karin Jacobs, initially had something quite different in mind. Originally, the team set out to research and describe the characteristics of hydrophobins – a group of naturally occurring proteins. ‘We noticed that the hydrophobins form colonies when they are placed in water. They immediately arrange themselves into tightly packed structures at the interface between water and glass or between water and air,’ explains Karin Jacobs. ‘There must therefore be an attractive force acting between the individual hydrophobin molecules, otherwise they would not organize themselves into colonies.’ But Professor Jacobs, research scientist Dr Hendrik Hähl and their team did not know how strong this force was.

This is where the neighbouring research group led by Professor Ralf Seemann got involved. One of Seemann’s research teams, which is headed by Dr Jean-Baptiste Fleury, studies processes that occur at the interfaces between two liquids. The research team set up a minute experimental arrangement with four tiny intersecting flow channels, like a crossroads, and allowed a stream of oil to flow continuously from one side of the crossing to the other. From the other two side channels they injected ‘fingers’ of water which protruded into the crossing zone. As the hydrophobins tended to gather at the interface of the carrier medium, they were in this case arranged at the water-oil interface at the front of the fingers. The physicists then ‘pushed’ the two fingers closer and closer together in order to see when the attractive force took effect. ‘At some point the two aqueous fingers suddenly coalesced to form a single stable interface consisting of two layers,’ says Ralph Seemann. ‘The weird thing is that it also functions the other way around, that is, when we use oil fingers to interrupt a continuous flow of water,’ he explains. This finding is quite new, as up until now other molecules have only exhibited this sort of behaviour in the one or the other scenario. Normally proteins will orient themselves so that either their hydrophilic (‘water loving’) sides are in contact with the aqueous medium, or their hydrophobic (‘water fearing’) side is in contact with an oily medium. That a type of molecule can form stable bilayers in both environments is something wholly new.

Encouraged by these findings, the researchers decided to undertake a third phase of experiments to find out whether the stable bilayer could be reconfigured to form a small membrane-bound transport sac — a vesicle. They attempted to inflate the stable membrane bilayer in a manner similar to creating a soap bubble, but using water rather than air. The experiment worked. The cell-like sphere with the outer bilayer of natural proteins was stable. ‘That’s something no one else has achieved,’ says Jean-Baptiste Fleury, who carried out the successful experiments. Up until now it had only been possible to create monolayer membranes or vesicles from specially synthesized macromolecules. Vesicles made from a bilayer of naturally occurring proteins that can also be tailored for use in an aqueous or an oil-based environment are something quite new.

In subsequent work, the research scientists have also demonstrated that ion channels can be incorporated into these vesicles, allowing charged particles (ions) to be transported through the bilayer of hydrophobins in a manner identical to the way ions pass through the lipid bilayers of natural cells.

As a result, the physicists now have a basis for further research work, such as examining the means of achieving more precisely targeted drug delivery. In one potential scenario, the vesicles could be used to transport water-soluble molecules through an aqueous milieu or fat-soluble molecules through an oily environment. Dr Hendrik Hähl describes the method as follows: ‘Essentially we are throwing a vesicle “cape” over the drug molecule. And because the “cape” is composed of naturally occurring molecules, vesicles such as these have the potential to be used in the human body.’

The results of this research work were a surprise. Originally, the goal was simply to measure the energy associated with the agglomeration of the hydrophobin molecules when they form colonies. But the discovery that hydrophobin bilayers could be formed in both orientations, opened the door to experiments designed to see whether vesicles could be formed. That one thing would lead to another in this way, offers an excellent example of the benefits of this type of basic, curiosity-driven research. ‘The “discovery” of these vesicles is archetypal of this kind of fundamental research. Or to put it another way, if someone had said to us at the beginning: “Create these structures from a natural bilayer,” we very probably wouldn’t have succeeded,’ says Professor Karin Jacobs in summary.

The Astonishing Healthcare Tech of the Future Is Arriving

A group of Singularity University alum have created the first patented, real-time biosensing device. The tech lets you hack your health with biometrics by reading antioxidant level changes on the skin from the palm of your hand.

A mutation in a single DNA letter causes a painful and debilitating disease known as sickle-cell anaemia. Researchers have wrestled with this illness for more than 65 years, and have now added CRISPR–Cas9 gene editing to their armoury.

In a paper published 12 October in Science Translational Medicine1, researchers reported some success in correcting the mutation in mice, though they concede that human applications are still years away. The efficiency of the process is also slightly too low for practical use, cautions author Jacob Corn, a biochemist at the University of California, Berkeley.

But the advance — and other recent efforts — has given Corn hope that the first treatment to address the cause of sickle-cell anaemia could be only a few years away. “We now finally may have some paths to address the cause of the disease rather than the symptoms,” he says.

A single tweak
On paper, curing the disease seems almost straightforward: rewrite the mutated sequence in the blood-generating stem cells of the bone marrow so that it's normal, and a patient would, in theory, be free of the illness.

“It’s a single base change,” says Stuart Orkin, who studies blood diseases at the Dana-Farber Cancer Institute and the Boston Children’s Hospital in Massachusetts. “You’d think this would be the poster child of gene correction.”

But companies and academics alike have struggled to make progress, and even CRISPR–Cas9, for all of its ease and utility, is not yet up to the task.

Researchers can target the Cas9 enzyme — or other gene-editing enzymes — to cut specific sites of the genome, but what happens next depends on the natural DNA-repair processes in the cell. Most often, that repair causes small DNA deletions. A separate repair process inserts a new sequence at the cut site. But that correction occurs with vanishing frequency in the sleepy, infrequently dividing stem cells of the bone marrow that would be the target of a sickle-cell therapy.

Corn and his colleagues found a way to make the modification more efficient. They used a short DNA strand carrying the corrected sequence of the sickle-cell gene and engineered it to bind to the DNA strand left dangling after the Cas9 enzyme cuts. When used in cells taken from human sickle-cell patients, the technique increased the efficiency with which they could repair the defective genes by up to 25%.

Disappointing drop
The team then introduced these edited cells into lab mice. There, the efficiency of their technique plummeted: cells with a DNA deletion seemed to proliferate better in mice than cells with the corrected form of the gene. In mice, only up to 5% of the transplanted, edited cells produced the normal form of haemoglobin.

This is right at the threshold of what might be required to alleviate patients’ distress, says Corn. He adds that some physicians who have seen his data said they would be willing to try the approach in clinical trials. But Corn argues that it would be best to improve efficiency before it is deployed in people.

The refined technique will be a boon for researchers looking at other applications of gene editing, says Michael Holmes, vice-president of research at Sangamo Biosciences in Richmond, California. But it also highlights another problem: the attempts at gene editing leave plenty of genes with a deletion where the Cas9 enzyme cut. Some of those deletions could result in abnormal haemoglobin production — causing another serious condition called β-thalassaemia.

Sangamo also ran up against this problem, Holmes says, and decided to try a different approach. The company, and several others in the field, are using gene editing to disrupt expression of a gene that suppress the production of fetal-haemoglobin: a form of haemaglobin that is expressed in the developing fetus and resists sickling. Boosting the production of fetal-haemoglobin could thereby reduce the amount of sickle-shaped haemoglobin in adults.

The latest results with CRISPR–Cas9 support the idea that targeting fetal haemoglobin may still be the safer appraoch, says Holmes. “There's still a long way to go towards finding ways to safely correct the gene.”

Nature doi:10.1038/nature.2016.20782