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Fig. 2 | Molecular Neurodegeneration

Fig. 2

From: LRRK2 phosphorylates pre-synaptic N-ethylmaleimide sensitive fusion (NSF) protein enhancing its ATPase activity and SNARE complex disassembling rate

Fig. 2

LRRK2 interacts with NSF. a Extracts of purified cortical synaptosomes were incubated with anti-LRRK2 antibodies or rabbit IgG. The immunocomplexes were sedimented with protein G-Sepharose and the samples were resolved by SDS-PAGE and analyzed by immunoblotting with anti NSF and anti LRRK2 antibodies. Immunoblotting against synaptotagmin 1 (S-tagmin1), synaptophysin (S-physin) and synaptobrevin (S-brevin) were performed to confirm purity of synaptosomal preparation. b Flag-NSF full-length or domains (N, D1, D2) purified from HEK293T and bound to M2 flag resin were incubated with a mouse brain lysate. Samples were subjected to immunoblotting using anti-LRRK2 (MJFF2) or stained with Coomassie to show flag inputs. c Size exclusion chromatography fractions of HEK293T expressing ectopic flag-NSF alone or together with 2xMyc-LRRK2 spotted onto nitrocellulose membrane and probed with anti-flag antibody (n = 3 independent experiments). d Immunofluorescence of primary cortical neurons stained for endogenous LRRK2 and endogenous NSF (scale bar is 10 μm)

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Molecular Neurodegeneration

ISSN: 1750-1326