Catalog

ASAP is committed to accelerating the pace of discovery and informing a path to a cure for Parkinson’s disease through collaboration, research-enabling resources, and data sharing. We’ve created this catalog to showcase the research outputs and tools developed by ASAP-funded programs.

Protocol

LRRK2 RCKW protein purification

Protein purification protocol for tag-less LRRK2RCKW as done by Leschziner and Reck-Peterson Labs. Same protocol can be used to purify LRRK1RCKW as well. Original protocol by David Snead. Modified by Yu Xuan Lin and Mariusz Matyszewski for publication.

Protocol

Rab8a expression and purification

Recombinant Rab8a expression and purification protocol as used by the Leschziner and Reck-Peterson Labs.

Article

PKC isoforms activate LRRK1 kinase by phosphorylating conserved residues (Ser1064, Ser1074, and Thr1075) within the CORB GTPase domain

Leucine-rich-repeat-kinase 1 (LRRK1) and its homologue LRRK2 are multidomain kinases possessing a ROC-CORA-CORB containing GTPase domain and phosphorylate distinct Rab proteins. LRRK1 loss of function mutations cause the bone disorder osteosclerotic metaphyseal dysplasia, whereas LRRK2 missense mutations that enhance kinase activity cause PD. Here, the authors study the mechanism controlling LRRK1 activity and reveal a novel unexpected activation mechanism. View preprint.

Article

Published: PKC isoforms activate LRRK1 kinase by phosphorylating conserved residues (Ser1064, Ser1074, and Thr1075) within the CORB GTPase domain

Published: This study describes how PKC isoforms are able to phosphorylate LRRK1 at three sites in a key regulatory domain of the protein (GTPase domain) inducing LRRK1’s kinase activity. Interestingly, this is not seen with the PD-associated LRRK2, suggesting that PKC isoforms do not regulate LRRK2. View original preprint.

Protocol

Preparation of LRRK2 RCKW cryo-EM grids

This is Leschziner’s Lab updated protocol for making cryo-EM grids for LRRK2 RCKW. This protocol, when using lower protein concentration, results in better monomer and dimer formation than the old protocol.

Article

Structural basis for Parkinson’s Disease-linked LRRK2’s binding to microtubules

Preprint: LRRK2 mutations are a common cause of familial PD. In some circumstances, LRRK2 co-localizes with microtubules. The authors report a cryo-electron microscopy structure of the catalytic half of LRRK2, containing its kinase (closed conformation) and GTPase domains, bound to microtubules. Further, they identified amino acids that mediate microtubule binding.

Article

Structural Biology of LRRK2 and its Interaction with Microtubules

Review: This review focuses on new insights into the stucture of LRRK2’s cytosolic and microtubule-bound forms and challenges going forward.

Protocol

LRRK2RCKW Widefield fluorescence microtubule binding assay

This assay uses TMR labeled LRRK2 or LRRK1 RCKW to measure binding to microtubules in vitro