NMR studies of interactions between Bax and BH3 domain-containing peptides in the absence and presence of CHAPS

• BaxΔC interacts with CHAPS directly, but relatively weakly.
• BaxΔC interacts with BaxBH3 or BimBH3 peptide directly, and relatively strongly.
• Binding with BaxBH3 or BimBH3 resulted in substantial spectral changes to BaxΔC.
• BaxBH3 or BimBH3 bounded BaxΔC is monomeric, but dimerizes when CHAPS is presented.

Activation and oligomerisation of Bax, a key pro-apoptotic Bcl-2 family protein, are key steps in the mitochondrial pathway to apoptosis. The signals for apoptosis are conveyed by the distantly related BH3-only proteins, which use their short BH3 domain, an amphipathic α-helix, to interact with other Bcl-2 family members. Here we report an NMR study of interactions between BaxΔC and BH3 domain-containing peptides in the absence and presence of CHAPS, a zwitterionic detergent. We find for the first time that CHAPS interacts weakly with BaxΔC (fast exchange on the NMR chemical shift timescale), at concentrations below micelle formation and with an estimated Kd in the tens of mM. Direct and relatively strong-interactions (slow exchange on the NMR chemical shift timescale) were also observed for BaxΔC with BaxBH3 (estimated Kd of circa 150 μM) or BimBH3 in the absence of CHAPS. The interaction with either peptide alone induced widespread chemical shift perturbations to BaxΔC in solution which implies that BaxΔC might have undergone significant conformation change upon binding the BH3 peptide. However, BaxΔC remained monomeric upon binding either CHAPS or a BH3 peptide alone, but the presence of both provoked it to form a dimer.

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