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Figure 2 | Molecular Brain

Figure 2

From: Ca2+/Calmodulin-dependent protein Kinase II interacts with group I Metabotropic Glutamate and facilitates Receptor Endocytosis and ERK1/2 signaling: role of β-Amyloid

Figure 2

Purified GST-fusion proteins encoding the mGluR1/5 IL2 loop domain interact with GFP-CaMKIIα. A) Shown is a representative immunoblot showing the co-immunoprecipitation of G protein-coupled receptor kinase 2 and GFP-CaMKIIα with a GST mGluR1/5 IL2 domain fusion protein. 1 μg of GST protein was incubated with 500 μg of HEK293 cell lysates over-expressing either GRK2 or GFP-CaMKIIα. The immunoblots are representative of 3 independent experiments. B) Upper panel, shows the schematic representation of GST-IL2 fusion protein alanine scanning mutants previously used to identify the IL2 residues required for GRK2 binding to Group I mGluRs [39]. Middle panel shows a representative immunoblot showing the co-immunoprecipitation of GFP-CaMKIIα with the GST-IL2 fusion protein alanine scanning mutants and the expression of the GST fusions. Lower panel shows the densitometric analysis of the relative co-immunoprecipitation of GFP-CaMKIIα with the GST-IL2 fusion protein alanine scanning mutants compared to the wild-type IL2 GST fusion protein. The data represents the mean ± SD of 3 independent experiments. C) Representative immunoblots showing GFP-CaMKIIα co-immunoprecipitated FL-mGluR1b and FL-mGluR1b mutants (K691A and K692A) that do not bind GRK2. Also shown are FL-mGluR1a immunoprecipitates and GFP-CaMKIIα expression in cell lysates. Bar graph shows the relative co-immunoprecipitation of GFP-CaMKIIα with the FL-mGluR1b and FL-mGluR1b mutants (K691A and K692A). Data shown represents the means ± SD of three independent experiments.

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