Combined effort
Scientists at l' École Polytechnique Fédérale de Lausanne (EPFL) have successfully combined infrared spectroscopy (IR) and atomic force microscopy (AFM) in nanoIR to distinguish between disease-causing aggregation of proteins and normal activity, the findings could lead to new clues regarding neurodegenerative diseases.
The misfolding and aggregation of errant proteins in the brain seems to be the underyling biochemical problem that damages neurones and gives rise to the symptoms of neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's diseases. Now, EPFL scientists have been able to distinguish between the disease-causing aggregation forms of such proteins, which could lead to new approaches to drug discovery in neurodegenerative diseases.
The aggregation process in brain and spinal cord progresses through different forms, Before aggregating, single proteins misfold into anti-parallel or criss-crossed weave patterns. The misfolded proteins then begin to aggregate through intermediate form, ultimately growing into large stringy tangles known as fibrils. By this point, the aggregation has become lethal to the cells. Unfortunately, current imaging techniques cannot distinguish between these intermediate species and the final harmful fibrils.
Aggregation, that's the name of the game
The EPFL team, led by Giovanni Dietler, has now used infrared nanospectroscopy, or nanoIR in which AFM reveals the signature of the three-dimensional structure of each aggregation form, as well as its physical properties, such as stiffness and the IR component shows the subtle changes that take place in the protein's structure that cause and drive its aggregation.
"These two techniques are very useful," explains Dietler. "But individually, they cannot pinpoint the moment that the protein begins to misfold or find how the protein's structural properties relate to the structure of a particular aggregation form." Both elements of nanoIR have proved necessary to allow researchers to differentiate aggregation species.
The team has focused on the protein ataxin-3 in this work. When it mutates, ataxin-3 begins to aggregate and form fibrils with devastating consequences on motor control and coordination leading to the neurodegenerative disease spinocerebellar ataxia.
Monitoring
Using nanoIR, the researchers were able to monitor the evolution of individual ataxin-3 proteins as they aggregated. They looked at the stiffness of individual aggregated forms and then linked it to the number of weave patterns that they contained, providing a correlation between the two factors. This is the first demonstration of such research for individual aggregation forms. The team's findings regarding spinocerebellar ataxia were published in the journal Nature Communications and have an intriguing twist that might mean the results could also be extended to other diseases. The nanoIR approach showed that ataxin-3 misfolds after it aggregates, not before as one might expect from current theories on aggregation. In fact, the aggregation of ataxin-3 seems to begin with the individual protein, and then moves on to the formation of an intermediate with the original protein structure rather than a misfolded one. This could have significant implications for medicine as it at last confirms previous theories about protein misfolding that could not be tested with earlier techniques. Moreover, Dietler suggests that nanoIR has much greater potential yet in understanding protein aggregation in a range of diseases.
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