Parkinson's at nanoscale
The protein alpha-Synuclein plays an important role in Parkinson's disease. But how it works and what it does to the nerve cells is still largely a mystery. Biophysicist Martina Huber uses electron spin resonance to reveal new information about how the protein interacts with the nerve membrane: not all parts of the protein bind equally well; it depends on the membrane's electrical charge.
Photo: Martina Huber
Protein plaques
The protein αS is the main constituent of the protein plaques - the Lewy bodies - which are characteristic for Parkinson's. And in the hereditary variant of Parkinson's, which appears at a younger age than the much more frequent age-related variant, the gene that codes for the aS protein seems to have mutated. This gene defect causes the proteins to coagulate more readily.
Interaction with nerve membrane
The structure of the protein still remains largely unexplained, as does the interaction of the protein with the nerve cell membrane. This interaction is probably crucial to the biological function of the protein and for the formation of Lewy bodies.
Three different forms
αS is a small protein, with 140 amino acids. It is a fascinating protein, according to Huber. When it is dissolved in a solution, it has hardly any structure. But when it binds to the membrane, it takes on a clear structure, namely that of two linked helices. In plaques, the protein adopts a different structure, known as a beta sheet.
Horseshoe-shaped
In June of this year, Huber and her colleagues demonstrated (in the journal JACS) that the helix form tends to fold itself in a horseshoe shape over the membrane. This week in the journal ChemBioChem, they reveal new information about the protein, this time about its interaction with the nerve membrane. Huber and her group carried out this research together with colleagues from the University of Twente, with a group headed by V. Subramaniam.
Image: Earlier this year Huber and her colleagues discovered that the protein aS forms a horseshoe shape on binding to the membrane. (Image: FOM)
Binding to the membrane
Huber studies the protein at nanoscale, using electron spin resonance. This time she is looking at the affinity between membrane and protein, by using electron spin resonance to measure the speed of movement of labelled locations on the protein. And she discovered that not all locations on the protein bind equally well to the - artificial - membrane.
Greater affinity
If a membrane itself has a largely negative charge, the whole protein - two linked helix strings - bind to it, but as the charge becomes more neutral, part of the protein detaches from the membrane. That part of the protein must therefore be less positively charged than the rest, and consequently needs the membrane to exert a greater attraction.
Information transfer
Researchers believe that the interaction of the protein with the membrane has a strong influence on the likelihood of a person becoming ill. 'If too much protein binds to the membrane, the membrane could start to leak, for example,' according to Huber. 'We are dealing here with complex synaptic membranes, which are important in information transfer. So it's easy to imagine all kinds of things that could go wrong if that were to happen.
Debate on plaque formation
Plaque formation is most probably also related to the way in which the protein, in helix form, binds to the membrane. But how exactly? It's a subject researchers are still discussing. Huber: 'Some scientists believe that binding to the membrane encourages plaque formation, and on the other hand there are others who believe that proteins coagulate less quickly if they are bound to a membrane. But both are probably possible,' she says, now that she has discovered that not all parts of the protein bind equally well to the membrane. 'It depends on the conditions.'
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Electron spin resonance
Huber studies αS proteins which are linked to Parkinson's, using electron spin resonance. Electron spin resonance (ESR) is closely related to nuclear magnetic resonance (NMR). The Leiden physicists are the only scientists in the Netherlands to work on the development and refinement of ESR, which requires in-depth knowledge of instrumentation and spins. Huber: 'We replace one or more amino acids on a protein with so-called spin labels. We manipulate these using microwaves, whose signals we then collect.'
Using ESR you can measure the distances between two spin labels on a scale of half to - under ideal conditions - 8 nanometers, with a margin of error of 0.01 nm. The speed of movement of the rotating spins can be measured in nanoseconds. In a solution, the spin labels can rotate freely, but this freedom of movement decreases as they bond more to a membrane. Huber used an artificial membrane which is as similar as possible to a biological membrane. By mixing neutral and negatively charged lipids in varying quantities, she was able to give the membranes a different charge density.
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Biochemical research
Biochemists were already aware that αS proteins bind well to negatively charged membranes. Huber: 'But using biochemical techniques only allows you to say something about the protein as a whole: it binds well or it doesn't bind. We have been able to study it in much more detail.'
Ill
What does her discovery mean for the understanding of Parkinson's disease? Huber: 'For me as a biophysicist, how Parkinson's works in the human body is too far removed from my field of expertise. Synaptic membranes, for example, have a very complex structure, that you obviously don't have with my artificial membrane. But the major questions for me are: does the charge of a membrane alter when a person becomes ill? Can the protein, if it binds completely, enter the membrane more easily? And, if the membrane has a weaker charge, does the protein bind more readily to other proteins?
Spin-Label EPR on α-Synuclein Reveals Differences in the Membrane Binding Affinity of the Two Antiparallel Helices (p NA)
Malte Drescher, Frans Godschalk, Gertjan Veldhuis, Bart D. van Rooijen, Vinod Subramaniam, Martina Huber
ChemBioChem 26 September 2008 (advance of print)
Abstract
The research led by Martina Huber is part of the FOM Biomolecular Physics programme: project 03BMP03