Identification of an activation site in Bak and mitochondrial Bax triggered by antibodies

Abstract

During apoptosis, Bak and Bax are activated by BH3-only proteins binding to the α2–α5 hydrophobic groove; Bax is also activated via a rear pocket. Here we report that antibodies can directly activate Bak and mitochondrial Bax by binding to the α1–α2 loop. A monoclonal antibody (clone 7D10) binds close to α1 in non-activated Bak to induce conformational change, oligomerization, and cytochrome c release. Anti-FLAG antibodies also activate Bak containing a FLAG epitope close to α1. An antibody (clone 3C10) to the Bax α1–α2 loop activates mitochondrial Bax, but blocks translocation of cytosolic Bax. Tethers within Bak show that 7D10 binding directly extricates α1; a structural model of the 7D10 Fab bound to Bak reveals the formation of a cavity under α1. Our identification of the α1–α2 loop as an activation site in Bak paves the way to develop intrabodies or small molecules that directly and selectively regulate these proteins.

Lymphangiogenesis in Classical Hodgkin Lymphoma - Preliminary Study with Clinicopathological Correlations

“…It seems the overall level distribution of all the components of the BCL2 family is of more significance than the status of each individual member of the family. The activation of the different members may also be critical 29. The association of the tumor cells apoptosis with lymphangiogenesis, although not immediate, is clearly present, if one recapitulates the results of the pilot study.…”

Direct Activation of BAX by BTSA1 Overcomes Apoptosis Resistance in Acute Myeloid Leukemia

“…In contrast, unbound BAX kept the α1–α2 loop in closed conformation during the MD trajectory (Figure S1E–S1G). Consistently, antibodies that either bind to or stabilize the closed conformation of α1–α2 loop of BAX were found to inhibit cytosolic BAX (Iyer et al, 2016, Uchime et al, 2016). Taken together, BTSA1 binds with high affinity to BAX at a critical region associated with BAX activation, which is consistent with the high binding selectivity for BAX.…”

Progress in targeting the BCL-2 family of proteins

“…However, small molecules targeting this canonical BH3-groove are still elusive. Interestingly, Kluck and colleagues identified an antibody (7D10), which bound to the α1–α2 loop of BAK, opposite to the canonical BH3-groove and promoted BAK conformational activation and oligomerization leading to MOMP [64]. The physiological relevance of this novel activation mechanism has not yet been established, although a movement of this loop on BAX to open position was determined to be essential upon BH3 trigger site activation [53].…”