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. 2009 Jan 30;10(3):285–292. doi: 10.1038/embor.2008.246

Figure 3.

Figure 3

Physical interaction between DAPK and beclin 1. (Aa) Protein extracts from COS7 cells, HEK293T cells, or no extracts were added to bacterially produced GST or GST–beclin-1. The pulled-down proteins, as well as the total cell extracts, were blotted with the indicated antibodies. (Ab) Ponceau S staining of GST and GST–beclin-1 to which the extracts were added. (B) HEK293 cells were co-transfected with Flag-tagged beclin 1 or with Flag-tagged beclin 1 lacking the Bcl-2-binding domain (ΔBD) together with Bcl-XL and HA-tagged DAPK. Beclin 1 was immunoprecipitated using Flag antibodies, and the co-immunoprecipitated proteins, as well as the total cell extracts, were blotted with DAPK, Bcl-XL and beclin 1 antibodies. (C) HEK293 cells were transfected with Flag–beclin-1 or Flag–GFP, and the Flag-tagged proteins were immunoprecipitated using Flag antibodies, and eluted with an excess of Flag peptides. The blot was reacted with DAPK antibodies or with Flag antibodies at different time exposures. DAPK, death-associated protein kinase; GFP, green fluorescent protein; GST, glutathione S-transferase; HA, haemagglutinin; HEK, human embryonic kidney.