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

The Biochemical Journal
James A IrvingDavid A Lomas

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 α1-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 mAb5E3 does not bind to the native state of α1-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 mAb5E3 occurs without affecting the energetic barrier to polymerization. As mAb5E3 also does not alter the relative stability of the monomer to intermediate, it acts in a manner similar to the E34...Continue Reading

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Citations

Jun 20, 2015·American Journal of Respiratory Cell and Molecular Biology·Imran HaqDavid A Lomas
Apr 2, 2016·Journal of Hepatology·David A LomasBibek Gooptu
May 26, 2016·Proceedings of the National Academy of Sciences of the United States of America·Kshama ChandrasekharDaniel N Hebert
Jul 14, 2016·The Biochemical Journal·Neda Motamedi-ShadDavid A Lomas
Dec 29, 2018·Monoclonal Antibodies in Immunodiagnosis and Immunotherapy·Fatemeh MalaeiFatemeh Rahbarizadeh
Jun 3, 2021·Materials·Samuele RaccostaMauro Manno
Jun 3, 2021·International Journal of Molecular Sciences·Riccardo RonzoniAnnamaria Fra

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Methods Mentioned

BETA
ELISA
FRET
PCR
electrophoresis
thermal shift
NMR

Software Mentioned

PyMOL
Prism GraphPad

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