Heat-induced dimerization of BCL-xL through α-helix swapping

The dimerization of anti-apoptotic BCL-x(L) by three-dimensional domain swapping has recently been discovered at alkaline pH; however, the high energetic barrier between the dimer and monomer forms of BCL-x(L) prevents them from interconverting at room temperature and neutral pH. Here, we demonstrate that BCL-x(L) dimers can be easily prepared by heating concentrated protein above 50 degrees C. The 38 kDa BCL-x(L) dimer was fully characterized by multi-resonance nuclear magnetic resonance (NMR) spectroscopy, and the mechanism of dimerization by alpha-helix swapping was confirmed. Dimerization strongly affects the NMR signals from the turn between helices alpha 5 and alpha 6 of BCL-x(L) and a portion of the long loop between helices alpha 1 and alpha 2. Measurements of residual dipolar couplings demonstrate that the solution structure of the BCL-x(L) dimer is very close to the crystal structure. Dimer formation does not prevent tight binding of ligands to the hydrophobic cleft of BCL-x(L); however, binding of a BID BH3-peptide or a polyphenol drug, gossypol, to BCL-x(L) significantly slowed monomer-dimer interconversion and is an example of the control of BCL protein oligomerization by ligand binding.