Investigation of the mechanical properties of filamentous protein aggregates using optical tweezers.

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Protein molecules carry out many if not most of the essential functions of life. However very occasionally, and only under abnormal conditions, many proteins aggregate into linear filaments containing thousands of copies of the same protein molecule but in an essentially useless form. These rope-like structures, known as fibrils , are typically one-10,000th the size of a human hair. The process of fibril formation has been the focus of intensive research because of its occurrence in many health disorders such as Alzheimer's Disease, Parkinson's Disease and, most famously, Mad Cow Disease. In particular it has been suggested that the mechanical properties of fibrils might play a role in some of these pathologies. For certain, a complete understanding of the filament growth process is lacking and would be of great value. Previous work in this area has explored the properties of protein filaments with a range of techniques able to access nanometre length-scales, including electron microscopy and atomic force microscopy (AFM), and this has led to a good understanding of the structural arrangement of the protein units. The elastic properties of some filaments have also been measured with AFM after binding to a solid support. However, the existing approaches suffer from a number of limitations and the challenge is to achieve a model experiment that might mimic conditions in the human body more closely. In particular we have in mind to reduce the interaction with synthetic surfaces and to measure directly the phenomenally tiny forces that act on the single filament level during growth. We therefore propose to study the growth and mechanical properties of protein fibrils using an optical tweezer apparatus. Objects that bend light more than the liquid in which they are suspended (for instance, a very small polystyrene particle suspended in water) can be trapped by the pressure of the photons present in a laser beam. The typical trapping forces on a particle are exceptionally small - between 0.1 and 100 piconewtons, less than one hundredth of a millionth of a millionth of the force required to kick a football - and the position of the particle can be determined down to a few nanometers. By simply steering the laser beam we can then move these trapped objects. This is the principle behind optical tweezers. The equipment we intend to use will enable us to control the position and direction of a growing protein fibril, keeping it away from any interfering surface. By causing a fibril to wiggle in a defined way, we hope to observe a phenomenon known as one-armed swimming , where the filament propels itself along. Furthermore, by pointing a growing fibril into a wall and pushing the free end with our laser beam, we will test whether an external structure will stop ( stall ) the fibril growth. From the biophysical point of view the novelty of this study lies in investigating a medically important system. From the physical point of view, we hope that assembling a range of filaments with different sizes and protein composition will enable us to develop a palette of filaments with a wider range of mechanical properties. These will be used more generally to investigate the universal behaviour of semi-flexible filaments.