An antibody raised against a pathogenic serpin variant induces mutant-like behaviour in the wild-type protein

Abstract

A monoclonal antibody (mAb) that binds to a transient intermediate may act as a catalyst for the corresponding reaction; here we show this principle can extend on a macro molecular scale to the induction of mutant-like oligomerization in a wild-type protein. Using the common pathogenic E342K (Z) variant of α ₁-antitrypsin as antigen–whose native state is susceptible to the formation of a proto-oligomeric intermediate–we have produced a mAb (5E3) that increases the rate of oligomerization of the wild-type (M) variant. Employing ELISA, gel shift, thermal stability and FRET time-course experiments, we show that mAb _5E3 does not bind to the native state of α ₁-antitrypsin, but recognizes a cryptic epitope in the vicinity of the post-helix A loop and strand 4C that is revealed upon transition to the polymerization intermediate, and which persists in the ensuing oligomer. This epitope is not shared by loop-inserted monomeric conformations. We show the increased amenity to polymerization by either the pathogenic E342K mutation or the binding of mAb _5E3 occurs without affecting the energetic barrier to polymerization. As mAb _5E3 also does not alter the relative stability of the monomer to intermediate, it acts in a manner similar to the E342K mutant, by facilitating the conformational interchange between these two states.

Abstract

We show that a monoclonal antibody can act as a ‘molecular template’ in aberrant protein oligomerization, and the transient intermediate of α ₁-antitrypsin, a key to the molecular mechanism of disease pathogenesis, expresses a cryptic epitope also present in the oligomer.