Figure 2

N-terminus of AnxA6 mediates binding with α-actinin. A. Scheme representing AnxA6 deletion mutants. The full length AnxA6 contains eight repeat domains and associated linker residues (residues 1-673). Deletion mutants GST-AnxA6 ΔN1 lacks N terminal tail and repeat domain 1 (residues 1-89); GST-AnxA6 ΔN2 lacks N terminal tail, repeat domains N1 and N2 (residues 1-163); GST-AnxA6 ΔN3 lacks N terminal tail, repeat domains N1, N2 and N3 (residues 1-250); GST-AnxA6 ΔC1 lacks C terminal tail and repeat domain C1 (residues 600-673); GST-AnxA6 ΔC3 lacks C terminal tail and repeat domains C1, C2 and C3 (residues 435-673). GFP-AnxA6 D N1 lacks N terminal tail and repeat domain 1 (residues 1-89) and GFP-AnxA6 D C1 lacks C terminal tail and repeat domain C1 (residues 600-673). B. Solubilised WHH was incubated with GST affinity beads (vector) or various mutants of AnxA6 lacking domains N1-N3 (GST-AnxA6 ΔN1- GST-AnxA6 ΔN3) or mutants lacking domains C1-C3 (GST-AnxA6 ΔC1 and GST-AnxA6 ΔC3). The presence of α-actinin in the bound fractions was detected by immunoblotting analysis with anti sarcomeric α-actinin antibody (arrow head).