Immunologic properties of enzymatically degraded human keratin intermediate filaments

In order to gain insight into the metabolism of keratin intermediate filaments (KIF) as well as the ability of KIF degradation products to interact with the immune system, we performed enzymatic degradation of purified KIF and examined their interaction with anti-KIF autoantibodies and their ability to act as immunogens. Aliquots of KIF aggregates were exposed to 3 different enzymes, that is, alpha-chymotrypsin, plasmin, and trypsin, in dose- and time-dependent experiments. The effect of the digestion was monitored sequentially by polyacrylamide gel electrophoresis and simultaneously by transmission electron microscopy. Furthermore, the KIF degradation proteins were then examined for their ability to bind anti-KIF autoantibodies by immunoblot and for their immunogenicity. In addition, preincubation of KIF with anti-KIF autoantibodies prior to the digestion procedure was performed to investigate a possible protective effect of this treatment against proteolytic degradation. The experiments demonstrated that: (1) KIF are degraded by serine proteinases, (2) with prolonged incubation time intact KIF are progressively replaced by more granular-amorphous material in transmission electron microscopy, (3) anti-KIF autoantibodies bind to KIF degradation proteins, (4) preincubation of KIF with anti-KIF autoantibodies does not exert any major protective effect for KIF against proteolytic degradation, and (5) the enzymatic degradation products of KIF can function as effective immunogens causing the formation of high-titer anti-KIF antibodies.