The conformational state of human alpha 2-macroglobulin influences its dissociation into half-molecules by sodium thiocyanate.
Sodium thiocyanate dissociates native human alpha 2-macroglobulin into half-molecules consisting of two disulfide-bonded subunits, when the salt concentration is equal to or exceeds 1.2 M. Incubation with 1.6 M sodium thiocyanate for 1 h at 22 degrees C dissociates about 90% of alpha 2-macroglobulin into half-molecules. The half-molecules remain stable when the concentration of sodium thiocyanate is reduced to 0.2 M or zero, demonstrating that reassociation does not occur under these conditions. The internal thiol esters of the half-molecules are intact because they can be exposed by treatment with methylamine or trypsin. The noncovalent interaction between the disulfide-bonded dimers is stronger in the "closed-trap" than in the "open-trap" conformation of alpha 2-macroglobulin. The cleavage in the bait region by trypsin makes alpha 2-macroglobulin completely stable toward dissociation, and alpha 2-macroglobulin remains in a tetrameric state in 2.2 M sodium thiocyanate even when trypsin is not covalently bound to it. The increase in fluorescence with time indicates that conformational changes occur as a consequence of dissociation.