Structures of proteins linked to Parkinson’s disease provide a framework for understanding how they work together

Scientists at St. Jude Children’s Research Hospital have revealed the complex structure of two proteins linked to Parkinson’s disease, both implicated in late-onset cases. Leucine-rich repeat kinase 2 (LRRK2) is a protein kinase that modifies other proteins in a process called phosphorylation; Rab29, a member of the Rab GTPase family that regulates cellular trafficking, modulates LRRK2 activity.

How Rab29 and LRRK2 act synergistically to cause Parkinson’s disease remains unclear. St. Jude researchers determined the structures of LRRK2 bound to Rab29, uncovering mysteries behind LRRK2 regulation and insights with implications for drug design.

The work was published today in Science.

Parkinson’s disease is the second most common neurodegenerative disease after Alzheimer’s disease and affects 1 to 2% of the population over the age of 65. The genetic link to the disease is well known, with around 15% of cases having a family history. Although there is a long list of genes associated with the disease, LRRK2 mutation is one of the most common causes. Due to its large size, structural studies on LRRK2 have been tedious.

“It is extremely difficult to work with this protein,” said corresponding author Ji Sun, Ph.D., Department of Structural Biology at St. Jude.

Despite these difficulties, Sun and his team presented the first structure of the complete LRRK2 in 2021 in Cell.

“In this first paper, we obtained the structure of LRRK2, but this structure showed an inactive conformation,” Sun explained. Proteins often have active and inactive forms, regulated by different cellular signals. Sometimes it takes binding to another protein to trigger the structural changes that move a protein from an inactive to an active form. “So we started thinking, ‘We have a key state of LRRK2.’ Can we get its active conformation?”

Cryo-electron microscopy captures the active state of LRRK2

Finding the active conformation was not as simple as adding Rab29 to LRRK2. LRRK2 can bind to other LRRK2 molecules in a process called oligomerization. This can transform a single LRRK2 monomer (one unit) into a dimer (two units) or even larger assemblies. This meant that researchers had to look for the version that represented the active form. There was also the problem that Rab29 was localized at cell membranes.

“In cells, about 90% or more of the cytosol LRRK2,” Sun explained. “A very small amount is located on the surface of the membrane and forms large oligomers. And it is these versions that are active and functional.”

Using cryo-electron microscopy, the researchers, including first author Hanwen Zhu, Ph.D., of St. Jude’s Department of Structural Biology, determined the first structures of the Rab29-LRRK2 complex. This included the structures of the monomer (one pair) and dimer (two pairs), but also an unexpected tetramer (four pairs).

“In this tetramer, we see the active conformation of LRRK2, but in the monomeric and dimeric complexes, LRRK2 is in an inactive conformation,” Sun said.

Understanding the Rab29-LRRK2 complex

These results demonstrate that LRRK2 is activated not only by the proteins with which it interacts, but also by their spatial arrangement within cells.

“We propose a transition from monomer to tetramer during membrane recruitment,” Sun explained. “Inside the cell, it is mostly inactive LRRK2 monomers or dimers. But when Rab29 recruits LRRK2 to the membrane, the local concentration of LRRK2 increases. This then facilitates the transition to the tetramer, in which LRRK2 becomes active.”

What are the implications for Parkinson’s disease? These structures provide researchers with an atomic-scale map to trace how the different mutations causing Parkinson’s disease affect the functioning of this complex.

“All of these mutations actually favor the active conformation, meaning they either provide new interactions within the active conformation or disrupt interactions within the inactive conformation,” Sun said. “The effects of mutations can be beautifully visualized in our structures; it is very well explained.”

The importance of such structural studies lies not only in the knowledge gained, but also in their potential application to drug design. For example, the researchers also captured the structure of LRRK2 in the presence of the drug DNL201. This drug, which was in a phase 1 clinical trial, locks the protein in an active state, so it was used to validate their findings that the tetramer was indeed the active form of the complex.

“We now have an inactive conformation and an active conformation, so we can monitor the transition from the inactive state to the active state,” Sun explained. “These structures provide much-needed information for medicinal chemists to design new inhibitors against LRRK2 for the treatment of Parkinson’s disease.”