Figure 1. Domain structure of HMGB1, cysteine residues of HMGB1 and oxidation, hcDNA.

(A) Domain structure of HMGB1 with two DNA binding domains, and polyanionic C-tail. (B) Schematic representation of cysteine residues of HMGB1 and formation of a disulphide bridge between Cys23 and Cys45. (C) Mild oxidation of HMGB1 in the presence of Cu²⁺ results in increased mobility in PAGE due to formation of an intramolecular disulphide bond by opposing Cys23 and Cys45, in agreement with MALDI-TOF mass spectrometry. Equal amounts (4 µg) of oxidized or reduced HMGB1 samples were loaded on the SDS-15%-polyacrylamide gel. Notice that oxidization of HMGB1 compromised Coomassie blue-staining of the protein in the gel. Arrow indicates electrophoretic mobility of HMGB1 dimer (formed due to an intermolecular cross-link of two HMGB1 molecules via the disulphide bridge). (D) Schematic drawing of hemicatenated DNA loops (hcDNA). hcDNA was created from a sequence containing a tract of poly(CA)·poly(TG) that can form a loop maintained at its base by hemicatenane, i.e. the junction of two DNA duplexes in which one of the strands of one duplex passes between the two strands of the other duplex [26], [42]. The drawing was kindly provided by François Strauss (National Museum of Natural History, Paris, France).