Fig. 2: The dual-specific aptamer did not recognize the inactive GDF11/8 complex.

A Unprocessed GDF8/11 (inactive) undergoes Furin cleavage to form the latent complex (inactive). TLD processing converts the latent complex into the triggered state (active), and dissociation of the remaining prodomain yields an unbound mature ligand (active). The binding of an antagonist renders the ligand inactive. B, C Anti-GDF8 prodomain (B), and anti-GDF11/8 mature ligand (C) western blot of the latent complex (Latent GDF8 and Latent GDF8P11M; GDF11 mature ligand with GDF8 prodomain), latent complex treated with BMP1 (Latent GDF8 / BMP1 and Latent GDF8P11M / BMP1), and latent complex incubated with the same digestion conditions but without BMP1 (Latent GDF8 / BMP1 and Latent GDF8P11M / no BMP1). Black labels are samples prior to the pull-down and arrows are following the aptamer pull-down. In (B), BMP1 processing of the latent complexes was confirmed via the detection of the ~20 kDa band (arrows). Non-pulled-down samples are shown in black, and the samples are pulled-down by aptamers below the “pull-down samples” labeling. Immunoblotting of latent complex is first used for anti-prodomain blotting, then stripped and reblotted for anti-mature ligand. All aptamer pull-downs were processed simultaneously. Experiments were independently repeated twice with similar results. D Schema of BMP1 processing required for aptamer binding. Source data are provided as a Source Data File.GDF11/8 ligand (monomer): 12.4 kDa, GDF11/8 ligand (dimer): 25 kDa, BMP1 processed prodomain: ~20 kDa, latent GDF11/8: ~40 kDa, mixture of unprocessed and latent GDF11/8: ~50 kDa. BMP1 = bone morphogenetic protein 1; TLD = Tolloid protease; spec = specific. Experiments performed in duplicate.