Active Recombinant Human LRRK2 (G2385R), GST-tagged

• Cat.No.: LRRK2-109H
• Product Overview: Recombinant human LRRK2 (G2385R) (968-end) was expressed by baculovirus in Sf9 insect cells using an N-terminal GST tag.

Specification

• Species: Human
• Source: Sf9 Cells
• Tag: GST
• Protein Length: 968-end a.a.
• Description: LRRK2 or leucine-rich repeat kinase is a protein with an ankryin repeat region, a leucine-rich repeat (LRR) domain, a kinase domain, a DFG-like motif, a RAS domain, a GTPase domain, a MLK-like domain, and a WD40 domain. Mutations in LRRK2 are the most frequent known cause of autosomal dominant and idiopathic Parkinson"s disease with prevalent mutations being found within the GTPase and kinase domains. LRRK2 cooperates with MET to promote efficient tumor cell growth and survival in various cancers. Down-regulation of LRRK2 in cultured tumor cells compromises MET activation and selectively reduces downstream MET signaling to mTOR and STAT3.
• Form: Recombinant protein stored in 50mM Tris-HCl, pH 7.5, 150mM NaCl, 10mM glutathione, 0.1mM EDTA, 0.25mM DTT, 0.1mM PMSF, 25% glycerol.
• Bio-activity: The specific activity of LRRK2 was determined to be 6 nmol /min/mg
• Molecular Mass: ~210 kDa
• Purity: >70% by densitometry
• Applications: Kinase Assay, Western Blot
• Storage: Store product at –70 centigrade. For optimal storage, aliquot target into smaller quantities after centrifugation and store at recommended temperature. For most favorable performance, avoid repeated handling and multiple freeze/thaw cycles.
• Concentration: 0.05 μg/μl
• Publications:
  G2385r and I2020T mutations increase LRRK2 GTPase activity (2016)

Gene Information

• Gene Name: LRRK2 leucine-rich repeat kinase 2 [ Homo sapiens ]
• Official Symbol: LRRK2
• Synonyms: LRRK2; leucine-rich repeat kinase 2; PARK8, Parkinson disease (autosomal dominant) 8; leucine-rich repeat serine/threonine-protein kinase 2; DKFZp434H2111; FLJ45829; RIPK7; ROCO2; augmented in rheumatoid arthritis 17; PARK8; AURA17; DARDARIN
• Gene ID: 120892
• mRNA Refseq: NM_198578
• Protein Refseq: NP_940980
• MIM: 609007
• UniProt ID: Q5S007
• Chromosome Location: 12q12
• Pathway: Parkinsons disease, organism-specific biosystem
• Function: ATP binding; GTP binding; GTP-dependent protein kinase activity; GTP-dependent protein kinase activity; GTPase activator activity; NOT GTPase activity; MAP kinase kinase activity; nucleotide binding; protein binding; protein homodimerization activity; protein kinase activity; protein serine/threonine kinase activity; tubulin binding

Citation

G2385R and I2020T Mutations Increase LRRK2 GTPase Activity

Journal: BioMed Research International    PubMed ID: 27314038    Data: 2022/12/14

Authors: Dong Hwan Ho, Jihoon Jang, Wongi Seol

Article Snippet:The myc-tagged LRRK2 WT, G2019S, R1441C, D1994A, and G2385R plasmids were described previously [ , ].The myc-tagged LRRK2 WT, G2019S, R1441C, D1994A, and G2385R plasmids were described previously [ , ].. The GST-ΔN (950-2527) LRRK2 WT, G2019S, R1441C, and D1994A recombinant proteins were purchased from Invitrogen (Carlsbad, CA, USA) and GST-ΔN LRRK2 I2020T and G2385R were obtained from Creative BioMart (Shirley, NY, USA).. Antibodies were purchased and used as follows.Antibodies were purchased and used as follows.

The kinase activities of LRRK2 I2020T and G2385R mutants ? .

GTPase assays of various LRRK2 proteins. (a) GTPase assays of the commercially available recombinant LRRK2 proteins. (b) GTPase assays using the immunoprecipitates of myc-tagged LRRK2 proteins transiently expressed in HEK 293T cells. The assay was carried out using a commercial GTPase assay kit. The relative GTPase activity results are shown after a statistical analysis of three independent experiments (top). Each GTPase activity value was divided by the quantity of protein used in the assay measured using GST (a) or myc (b) antibodies. Representative Western blots were shown (bottom). All values are relative to the WT and are expressed as the mean ± SEM. Vec: empty vector, WT: wild type, GS: G2019S, DA: D1994A, RC: R1441C, IT: I2020T, and GR: G2385R. ? p ≤ 0.05, ?? p ≤ 0.01, and ??? p ≤ 0.001.

In vitro kinase assays with commercially available recombinant LRRK2 proteins. The assays were carried out using purified recombinant His-Rab5B protein as the substrate. (a) LRRK2 and Rab5B phosphorylations were shown by autoradiography. The quantities of the LRRK2 and Rab5B proteins used are shown in the Western blot results with the GST and Rab5B antibodies below the corresponding autoradiograms. Relative Rab5B phosphorylation was calculated and is shown below the Rab5B Western blot. (b) Relative kinase activities of autophosphorylated LRRK2 after a statistical analysis of three independent experiments. All values are relative to the WT and are expressed as the mean ± SEM. —: no added LRRK2 protein, WT: wild type, GS: G2019S, DA: D1994A, RC: R1441C, IT: I2020T, and GR: G2385R. ? p ≤ 0.05 and ?? p ≤ 0.01.