Identification of single‐domain, Bax‐specific intrabodies that confer resistance to mammalian cells against oxidative‐stress‐induced apoptosis

Bax is a proapoptotic protein implicated in cell death involved in several neurodegenerative diseases. Intracellularly expressed antibody (Ab) fragments (intrabodies) inhibiting Bax function would have potential for developing therapeutics for the aforementioned diseases and can serve as research tools. We report identification, cloning, and functional characterization of several Bax-specific single-domain antibodies (sdAbs). These minimal size Ab fragments, which were isolated from a llama V(H)H phage display library by panning, inhibited Bax function in in vitro assays. Importantly, as intrabodies, these sdAbs, which were stably expressed in mammalian cells, were nontoxic to their host cells and rendered them highly resistant to oxidative-stress-induced apoptosis. The intrabodies prevented mitochondrial membrane potential collapse and apoptosis after oxidative stress in the host cells. These anti-Bax V(H)Hs could be used as tools for studying the role of Bax in oxidative-stress-induced apoptosis and for developing novel therapeutics for the degenerative diseases involving oxidative stress.