Fig 1: Purification and characterization of DDX43 KH-domain proteins.A, the schematic representation of four DDX43 KH-domain fragments. The KH domain and GRGG loop are indicated in red and helicase core domain in yellow. B, the SDS-PAGE analysis of recombinant DDX43 KH-domain proteins eluting from a Sephacryl S-100 HR column. C, the Western blot analysis for proteins shown in panel B with an anti-His antibody (SC-8036, Santa Cruz). D–E, the representative EMSA images of increasing DDX43 KH-domain protein (126 aa, 0–9.6 μM) binding with 0.5 nM of indicated DNA (D) and RNA (E) substrates. See Table S2 for substrate sequence. DNA is in black and RNA in gray. EMSA, electrophoretic mobility shift assay; HR, high-resolution.
Fig 2: Anti-SOD1olig antibody reacts with the interior of SOD1 structure. a The translated products of five exons in SOD1 and the three peptides, Pep 1, 2, and 3, are shown in schematic representation of the SOD1 primary structure. The ligands for copper and zinc ions are also shown and colored blue and red, respectively. His 63 (colored green) is a ligand for bridging both copper and zinc ions. b Identification of the epitope for anti-SOD1olig antibody (filled bars) was performed by indirect ELISA using the dissected peptides of SOD1 shown in (a). The peptides were prepared as a fusion protein with an N-terminal 6x His tagged GST. Almost equal amounts of peptides were examined, which was confirmed by ELISA using anti-His tag antibody (sc-8036, Santa Cruz Biotechnology) (open bars). The ELISA signal was represented as a ratio against that obtained using BSA. c The region covering the epitope of anti-SOD1olig antibody (Gly44 - Asn53) is shown red in the crystal structure of SOD1 (PDB ID: 2C9V). Copper (blue) and zinc (orange) ions are also indicated with the ligands
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