Chemoselective Staudinger-induced Michael-additions to antibodies to analyze protein homeostasis in C. elegans

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In this proposal a novel metal-free chemoselective conjugation strategy will be applied to modify antibodies for the analysis of protein-protein interactions of key relevance in the maintenance of protein homeostasis. In this joint effort the expertise of the Hackenberger laboratory in the engineering of bioorthogonal reactions and the Kirstein laboratory in protein homeostasis will be combined to identify novel chaperone complexes involved in protein disaggregation and their protein substrates in C. elegans. In addition, we aim to employ antibody drug conjugates to target modifiers of amyloid formation to Abeta formed fibrils in vivo. With the development of a Staudinger-induced Michael-addition for the chemoselective conjugation of cysteine residues in antibodies a new concept for the modification of proteins will be introduced. This novel technique enables the modular and metal-free coupling of two complex functional building blocks without the need of intermediate protecting group manipulations. Additionally, a phosphonamidate moiety is introduced by this method, which allows the direct incorporation of a light- or enzymatically cleavable bond in protein conjugates. This new synthetic tool will be applied to obtain functional antibodies for the identification of unknown protein substrates as well as interaction partners of specific chaperones and will furthermore be used in the design of light-cleavable antibody drug conjugates to target amyloid fibrils in living animal models for Alzheimer's disease that express Abeta1-42 peptides.