title: Ca2+, K+-regulated intramolecular crosslinking of S-100 protein via disulfide bond formation.
abstract: Reaction of the thiol reagent 5,5'-dithio-bis(2-nitrobenzoic acid) (Nbs2) with the brain-specific protein S-100 favours stabilization of the quaternary structure of the protein via disulfide bond formation. This process is modulated by those cations (Ca2+ and K+) which are known to affect the conformation of the protein. Ca2+ markedly favours the reaction of S-100 with Nbs2 but inhibits subsequent disulfide bond formation; K+, on the contrary, is much less effective in promoting interaction with Nbs2 but strongly stimulates disulfide bond formation. These findings are interpreted assuming that in presence of Ca2+ the three subunits forming the native S-100 protein have two cysteine residues exposed to the solvent but mismatched to form disulfides while in presence of K+ the sulphydryl groups are in a less accessible position to Nbs2 but suitable for S-S bond formation. Crosslinking of S-100 subunits is characterized by the appearance in dodecylsulphate electrophoresis of two very close protein bands having a molecular weight almost identical to that of the native, undenatured protein but not of higher or lower-molecular weight components. This finding, and the demonstration that both the crosslinked and native S-100 proteins have identical profiles when analyzed by sucrose density centrifugation or gel chromatography indicate that disulfide bond formation occurs among subunits of the same molecule.
mesh_terms: D000818:Animals; D001665:Binding Sites; D001921:Brain; D002118:Calcium; D002417:Cattle; D004220:Disulfides; D004228:Dithionitrobenzoic Acid; D006736:Horses; D007700:Kinetics; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D011188:Potassium; D011485:Protein Binding; D011489:Protein Denaturation; D009418:S100 Proteins
chemical_list: D004220:Disulfides; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D009418:S100 Proteins; D004228:Dithionitrobenzoic Acid; D011188:Potassium; D002118:Calcium