Structural insights into the GTP-driven monomerization and activation of a bacterial LRRK2 homolog using allosteric...

Area of Science:

• Biochemistry
• Structural Biology
• Neuroscience

Background:

• Mutations in human Leucine-Rich Repeat Kinase 2 (LRRK2) are a primary genetic cause of familial Parkinson's disease.
• LRRK2 possesses both GTPase and protein kinase activities; disease-associated mutations enhance kinase activity and potentially impair GTPase function.
• The interplay between LRRK2's catalytic domains and the regulatory role of its GTPase domain remain poorly understood, with a lack of structural data for activated states.

Purpose of the Study:

• To determine the structure of a Roco protein in its active, GTP-bound state.
• To elucidate the conformational changes associated with Roco protein activation.
• To understand the mechanism of allosteric activation by nanobodies and its relevance to LRRK2.

Main Methods:

• Single-particle cryo-electron microscopy (cryo-EM) was employed to resolve the structure.
• Two conformation-specific nanobodies, NbRoco1 and NbRoco2, were used as tools to stabilize and facilitate structural determination.
• A bacterial Roco protein (CtRoco) was utilized as a model system.

Main Results:

• The cryo-EM structure of CtRoco in its GTP-bound state revealed an active, monomeric conformation.
• A significant GTP-induced conformational change was observed, with the LRR-Roc linker acting as a hinge.
• The nanobodies NbRoco1 and NbRoco2 were shown to allosterically activate CtRoco through collaborative binding.

Conclusions:

• The study provides the first structural view of an activated Roco protein, detailing a large conformational rearrangement upon GTP binding.
• The findings illuminate the activation mechanism of Roco proteins and offer critical insights into the regulation of LRRK2.
• The identified allosteric activation pathway suggests potential therapeutic strategies for modulating LRRK2 GTPase activity.