The limited proteolysis of tumor necrosis factor-alpha.

The limited proteolysis of human recombinant TNF-alpha by trypsin yields two stable products resulting from cleavage after Arg6 and Arg44. In solution these two products remain associated together in a trimer with a Stokes' radius slightly greater than the radius of intact TNF-alpha and, therefore, could not be separated from each other under nondenaturing conditions. This limited digest retains at least 20% of the activity of the original TNF-alpha sample, and has a tertiary structure that is similar to that of the native protein by circular dichroism. On the other hand, incorrectly folded, inactive TNF-alpha undergoes extensive digestion following similar treatment with trypsin. These results indicate that the active form of TNF-alpha has a tight core structure which is maintained after N-terminal cleavage and removal.