Ayan Adhikari
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View article: Allosteric regulation of the Golgi-localized PPM1H phosphatase by Rab GTPases modulates LRRK2 substrate dephosphorylation in Parkinson’s disease
Allosteric regulation of the Golgi-localized PPM1H phosphatase by Rab GTPases modulates LRRK2 substrate dephosphorylation in Parkinson’s disease Open
PPM1H phosphatase reverses Parkinson's disease-associated, leucine-rich repeat kinase 2 (LRRK2)-mediated, Rab GTPase phosphorylation. We showed previously that PPM1H relies on an N-terminal amphipathic helix for Golgi membrane localization…
View article: Lysosomal glucocerebrosidase is needed for ciliary Hedgehog signaling: A convergent pathway contributing to Parkinson’s disease
Lysosomal glucocerebrosidase is needed for ciliary Hedgehog signaling: A convergent pathway contributing to Parkinson’s disease Open
Parkinson’s disease is characterized by loss of dopamine neurons that project to the dorsal striatum, and mutations in LRRK2 and GBA1 are the most common genetic causes of familial Parkinson’s disease. Previously, we showed that pathogenic…
View article: Sucrose Density Gradient Co-flotation of PPM1H and Rab10 v1
Sucrose Density Gradient Co-flotation of PPM1H and Rab10 v1 Open
Sucrose gradient co-flotation assays are employed to assess the potential binding of mNeon-PPM1H phosphatase to unphosphorylated Rab10. Recombinant mNeon-PPM1H and Rab10 are incubated with 50 nm-diameter liposomes for 30 min at 30°C. The m…
View article: End-product inhibition of the LRRK2-counteracting PPM1H phosphatase
End-product inhibition of the LRRK2-counteracting PPM1H phosphatase Open
PPM1H phosphatase reverses Parkinson’s disease-associated, Leucine Rich Repeat Kinase 2-mediated, Rab GTPase phosphorylation. We showed previously that PPM1H relies on an N-terminal amphipathic helix for Golgi membrane localization and thi…
View article: PPM1H Purification from E. coli v1
PPM1H Purification from E. coli v1 Open
This protocol is used to purify full length PPM1H enzyme in good yield and purity after expression in bacteria. We generally obtain 2mg from 4L bacterial culture and express the protein at 18°C to avoid aggregation due to overexpression.
View article: PPM1M, a LRRK2-counteracting, phosphoRab12-preferring phosphatase with potential link to Parkinson’s disease
PPM1M, a LRRK2-counteracting, phosphoRab12-preferring phosphatase with potential link to Parkinson’s disease Open
Leucine-rich repeat kinase 2 (LRRK2) phosphorylates a subset of Rab GTPases that regulate receptor trafficking; activating mutations in LRRK2 are linked to Parkinson’s disease. Rab phosphorylation is a transient event that can be reversed …
View article: Dephosphorylation of phosphorylated Rab GTPases by PPM phosphatases v1
Dephosphorylation of phosphorylated Rab GTPases by PPM phosphatases v1 Open
This protocol describes our method to measure phosphoRab GTPase dephosphorylation using purified phosphoRab GTPases and PPM family phosphatases. We include our protocol for generation of phosphatase substrates by phosphorylation of Rab GTP…
View article: Lysosomal Glucocerebrosidase is needed for ciliary Hedgehog signaling: A convergent pathway to Parkinson’s disease
Lysosomal Glucocerebrosidase is needed for ciliary Hedgehog signaling: A convergent pathway to Parkinson’s disease Open
SUMMARY Mutations in LRRK2 and GBA1 are the most common genetic causes of familial Parkinson’s disease. Previously, we showed that pathogenic LRRK2 mutations inhibit primary cilia formation in rare interneurons and astrocytes of the mouse …
View article: Detection of accessible cholesterol in primary cilia using purified His-ALOD4-mNeon in 3T3 Fibroblasts v1
Detection of accessible cholesterol in primary cilia using purified His-ALOD4-mNeon in 3T3 Fibroblasts v1 Open
There exist at least three different pools of cholesterol in the plasma membrane: the essential pool, the sphingomyelin-sequestered pool, and the accessible pool (Radhakrishnan et al, 2020). Ciliary Hedgehog signaling is regulated by the a…
View article: Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis
Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis Open
PPM1H phosphatase reverses Parkinson’s disease-associated, Leucine Rich Repeat Kinase 2-mediated Rab GTPase phosphorylation. We show here that PPM1H relies on an N-terminal amphipathic helix for Golgi localization. The amphipathic helix en…
View article: Preparation of unilamellar liposomes v1
Preparation of unilamellar liposomes v1 Open
We present here a protocol for preparing liposomes that can be used to monitor binding of proteins to vesicles of various membrane curvatures. We used this to study the association of the PPM1H phosphatase with highly curved membranes due …
View article: Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis
Localization of PPM1H phosphatase tunes Parkinson’s disease-linked LRRK2 kinase-mediated Rab GTPase phosphorylation and ciliogenesis Open
PPM1H phosphatase reverses Parkinson’s disease-associated, LRRK2-mediated Rab GTPase phosphorylation. We show here that PPM1H relies on an N-terminal amphipathic helix for Golgi localization. The amphipathic helix enables PPM1H to bind to …
View article: A feed-forward pathway drives LRRK2 kinase membrane recruitment and activation
A feed-forward pathway drives LRRK2 kinase membrane recruitment and activation Open
Activating mutations in the leucine-rich repeat kinase 2 (LRRK2) cause Parkinson’s disease, and previously we showed that activated LRRK2 phosphorylates a subset of Rab GTPases (Steger et al., 2017). Moreover, Golgi-associated Rab29 can re…
View article: Author response: A feed-forward pathway drives LRRK2 kinase membrane recruitment and activation
Author response: A feed-forward pathway drives LRRK2 kinase membrane recruitment and activation Open
Article Figures and data Abstract Editor's evaluation Introduction Results Discussion Materials and methods Data availability References Decision letter Author response Article and author information Metrics Abstract Activating mutations i…
View article: A Feed-forward Pathway Drives LRRK2 kinase Membrane Recruitment and Activation
A Feed-forward Pathway Drives LRRK2 kinase Membrane Recruitment and Activation Open
Activating mutations in the Leucine Rich Repeat Kinase 2 (LRRK2) cause Parkinson’s disease and previously we showed that activated LRRK2 phosphorylates a subset of Rab GTPases (Steger et al., 2017). Moreover, Golgi-associated Rab29 can rec…
View article: Reconstitution of LRRK2 membrane recruitment onto planar lipid bilayers v1
Reconstitution of LRRK2 membrane recruitment onto planar lipid bilayers v1 Open
Supported lipid bilayers have emerged as an ideal model system to study the interaction of proteins with cellular membranes. We describe here a method to monitor the recruitment of purified LRRK2 kinase onto planar lipid bilayers containin…
View article: Reconstitution of LRRK2 membrane recruitment onto planar lipid bilayers v1
Reconstitution of LRRK2 membrane recruitment onto planar lipid bilayers v1 Open
Supported lipid bilayers have emerged as an ideal model system to study the interaction of proteins with cellular membranes. We describe here a method to monitor the recruitment of purified LRRK2 kinase onto planar lipid bilayers containin…
View article: Regulation of <i>Leishmania major</i> PAS domain‐containing phosphoglycerate kinase by cofactor Mg<sup>2+</sup> ion at neutral pH
Regulation of <i>Leishmania major</i> PAS domain‐containing phosphoglycerate kinase by cofactor Mg<sup>2+</sup> ion at neutral pH Open
Recently, we described the PAS domain‐containing phosphoglycerate kinase (PGK) from Leishmania major (LmPAS‐PGK) that shows acidic pH (5.5)‐dependent optimum catalytic activity. The PAS domain of LmPAS‐PGK is expected to regulate PGK activ…