Mutations in one gene, LRRK2, are a common cause of Parkinson’s disease. However, the protein product of this gene is large and complex and, to date, no one has been able to make highly pure, functional LRRK2 that can be easily used for biochemical experiments. Our primary objective is to make purified protein by combining expertise in multiple technologies from each of our collaborating laboratories and to share our methods with other researchers. If successful, our secondary objective is then to perform an exhaustive search for other proteins that may interact with LRRK2.
In our first set of experiments, we will try and produce active LRRK2. Our first step will be to genetically engineer a vector system that is derived from viruses to express the gene that we are interested in. In this case, we will use full length LRRK2, which is associated with PD, and a comparitive protein, LRRK1, which is not. Next, we will use these viral particles to ‘trick’ cells in culture to making a great deal of the proteins that we want – we are particularly interested to see if we can maintain this over time so we can scale up the amounts of proteins that can be made. We will then use a series of purification steps so we can recover only the LRRK2 we want and not any other contaminants. We have a defined criteria for what we consider ‘successful’ and, if we can reach these criteria, we will make the appropriate cells and methods available to other scientists.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
If we can solve the problems that we currently know impede our production of pure intact and active LRRK2, we expect that this can be used as the first step in developing two things. First, we can look at how LRRK2 mutations influence neurons to cause disease by identifying which other proteins that it interacts. This may impact diagnosis by being a readout of ‘normal’ or ‘abnormal’ activity. In the longer term, we hope that it would also have impact on treatment as there are several examples now of therapies that were developed from having a well characterized protein on which to base a drug.
We expect that we will be able to determine if our combination of approaches will yield a helpful new way to make active LRRK2. Just as importantly, if our approach fails we will be able to share that knowledge. We also expect to obtain some fairly fundamental insights into how LRRK2 normally works, and what happens when we have a mutation in the gene that causes PD.