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. 2012 Feb 13;7(2):e31554. doi: 10.1371/journal.pone.0031554

Figure 1. Multiple domains in spinophilin directly bind purified CaMKII isoforms.

Figure 1

A. Stylized domain structure of spinophilin showing the F-actin-, G-protein coupled receptor-, and PP1-binding domains, as well as the coiled-coil domains. GST spinophilin fusion proteins containing the indicated fragments were generated. B. Non- or Thr287-autophosphorylated CaMKIIβ was incubated with GST, GSTSpN, or GSTSpC. Complexes were isolated using glutathione agarose and then immunoblotted for CaMKIIβ. C. Non- or Thr286-autophosphorylated CaMKIIα was incubated with GST or the indicated GST-fusion proteins. Complexes were isolated as in Panel B and immunoblotted for CaMKIIα. D. F-actin was incubated with GST, GSTSpN1, or GSTSpN2. Complexes were isolated using glutathione agarose and immunoblotted for actin. E. GSTSpN2 was incubated with CaMKII or F-actin in the presence of increasing concentrations of F-actin or CaMKII, respectively. Complexes were isolated using glutathione agarose and immunoblotted for CaMKII and actin. F. GSTSpN was incubated with CaMKIIα (250 nM) in the presence of increasing concentrations of F-actin. Complexes were isolated using glutathione agarose and immunoblotted for CaMKIIα and actin. All figures are representative of at least 2 experiments.