Oligomerization process of Bcl-2 associated X protein revealed from intermediate structures in solution

Abstract

Upon apoptotic stress, Bcl-2 associated X (BAX) protein undergoes conformational changes and oligomerizes, leading to the mitochondrial membrane permeabilization and cell death. While structures of the resultant oligomer have been extensively studied, little is known about the intermediates that describe the reaction pathway from the inactive monomers to activated oligomers. Here we characterize the intermediate structures of BAX using combined small-angle X-ray scattering (SAXS) with on-line gel-filtration and electron spin resonance (ESR). The intermediates, including monomers, dimers, and tetramers, are reconstructed via integrating the SAXS-envelopes and ESR-determined skeleton structures. The hence revealed structures suggest a linear oligomerization of BAX utilizing the extended dimers with the two flexible α6 chains protruded out as ditopic ligands. The results of molecular dynamics simulation also support the ditopic dimer conformation with mobile α6. The ditopic dimers could further wind into a helical rod structure with three dimers in one helical turn. Our results not only reveal the on-pathway intermediates, but also suggest a ditopic oligomerization mechanism that may bridge the observed intermediate structures in solution to the large BAX assemblies lately observed on mitochondria.

Overexpressed IGFBP5 promotes cell proliferation and inhibits apoptosis of nucleus pulposus derived from rats with disc degeneration through inactivating the ERK/MAPK axis

“…In addition, after transfection, the levels of Bax and caspase‐3 increased, meanwhile Bcl‐2 level decreased in the IGFBP5‐shRNA group, however, the opposite results were obtained from the IGFBP5 overexpression and PD98059 groups. In the apoptotic stress, Bax protein experienced the conformational changes and oligomerizes, leading to the cell death and mitochondrial membrane permeabilization . In addition to mitochondria, Bcl‐2 is situated at the endoplasmic reticulum in which it is a component of several distinguishing complexes .…”

“…In the apoptotic stress, Bax protein experienced the conformational changes and oligomerizes, leading to the cell death and mitochondrial membrane permeabilization. 39 In addition to mitochondria, Bcl-2 is situated at the endoplasmic reticulum in which it is a component of several distinguishing complexes. 40 NP cells were transfected with codon optimized Bcl-2 gene, commonly known to efficiently decline the serum starvation-induced cell apoptosis, and the overexpression of Bcl-2, while also decreasing the mRNA level of caspase-3.…”

Identifying Protein Conformational Dynamics Using Spin‐label ESR

“…Although it was ar ather time-consuming process, the iterative evaluation procedure was proven valuablet o derive ar easonably representative model for the activated BAX, in ag ood agreement with the structures reported later using other methodsi ncluding SAXS, cryo-EM, super-resolution fluorescencemicroscopy. [39][40][41][42] DEER technique has in recenty ears becomeagreat complement to other biophysical tools such as NMR andf luorescence techniques in revealing conformational heterogeneity of protein. [43][44][45] Aashish et al,f or example, utilized the 19 FNMR and DEER to study the conformation ensemble of the b2-Adrenergic receptor (b2AR).…”

Doubly spin-labeled nanodiscs to improve structural determination of membrane proteins by ESR

“…Clearly, the sensitivity of the DEER-derived results is improved, hence making it possible to discern the small changes in the ND geometry associated with the incorporation of YetJ. Potentially, the improved ability of PDS with ND + , when combined with small-angle X-ray scattering data, 7,28 can provide even more valuable information about molecular details of ND environment.…”