Overview
Cataract means that the lens at the front of the eye is slowing changing from clear to cloudy. As it changes, vision becomes blurred or hazy.
Age-related cataract is the cause of almost half of the world’s blindness. It happens when protein molecules in the lens stop working normally and start to build up.
Research has shown that this build-up begins when proteins ‘misfold’. Usually, proteins need to be folded into the right 3D shape in order to work.
In this project Dr Garcia-Manyes’ student will use a new technique known as ‘single molecule force-clamp spectroscopy’. They want to study each step taken by individual protein molecules as they unfold and refold. The aim is to pinpoint the exact step at which folding starts to go wrong and to track what happens afterwards.
Results from the project will help us understand, for the first time, how protein misfolding in the lens begins at the level of a single protein. This is a vital step towards developing non-surgical treatment to prevent or delay cataract.
-
Scientific summary
Unveiling the molecular mechanisms underlying the onset of gamma-D-crystallin aggregation studied by single molecule force-clamp spectroscopy
Age-related eye cataract, which results from aggregation of lens crystallins, is responsible for 48% of world blindness. A common feature of several diseases that are triggered by protein misfolding is the formation of aggregates that organize into fibers and plaques. Unfortunately, the presence of such large precipitates is normally the signature of a late stage in the evolution of the disease. Hence, the Holy Grail lies in interrupting the aggregation process from its initial stage. However, little is known about the molecular mechanisms that trigger the onset of these diseases.
There is now considerable evidence that in the vast majority of conformational diseases, the onset is a rare event that occurs at the single molecule level during the folding trajectory of a protein. Single molecule force-clamp spectroscopy allows, for the first time, capture of the conformational dynamics visited by a single protein during its journey to the folded state from highly extended states.
Here the team is using a combination of molecular biology engineering techniques and single molecule force-clamp spectroscopy, to map out the individual (un)folding pathways of γD-crystallin, allowing them to directly identify the molecular conformation that triggers the misfolding and aggregation reaction. They are complementing these observations with mesoscopic fluorescence experiments and classical aggregation experiments conducted in the bulk.
Altogether, this series of innovative and cross-disciplinary experiments provides a remarkable and timely opportunity to expand the current applications of force-clamp spectroscopy to solving the common riddle of the molecular origins of eye cataract, occurring at the single molecule level.