Bcl-2 and Bax interact via the BH1-3 groove-BH3 motif interface and a novel interface involving the BH4 motif.
The interaction of Bcl-2 family proteins at the mitochondrial outer membrane controls membrane permeability and thereby the apoptotic program. The anti-apoptotic protein Bcl-2 binds to the pro-apoptotic protein Bax to prevent Bax homo-oligomerization required for membrane permeabilization. Here, we used site-specific photocross-linking to map the surfaces of Bax and Bcl-2 that interact in the hetero-complex formed in a Triton X-100 micelle as a membrane surrogate. Heterodimer-specific photoadducts were detected from multiple sites in Bax and Bcl-2. Many of the interaction sites are located in the Bcl-2 homology 3 (BH3) region of Bax and the BH1-3 groove of Bcl-2 that likely form the BH3-BH1-3 groove interface. However, other interaction sites form a second interface that includes helix 6 of Bax and the BH4 region of Bcl-2. Loss-of-function mutations in the BH3 region of Bax and the BH1 region of Bcl-2 disrupted the BH3-BH1-3 interface, as expected. Surprisingly the second interface was also disrupted by these mutations. Similarly, a loss-of-function mutation in the BH4 region of Bcl-2 that forms part of the second interface also disrupted both interfaces. As expected, both kinds of mutation abolished Bcl-2-mediated inhibition of Bax oligomerization in detergent micelles. Therefore, Bcl-2 binds Bax through two interdependent interfaces to inhibit the pro-apoptotic oligomerization of Bax.
Ding
Jingzhen
J
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73126, USA.
Zhang
Zhi
Z
Roberts
G Jane
GJ
Falcone
Mina
M
Miao
Yiwei
Y
Shao
Yuanlong
Y
Zhang
Xuejun C
XC
Andrews
David W
DW
Lin
Jialing
J
D020816:Amino Acid Motifs; D000818:Animals; D006801:Humans; D009154:Mutation; D011485:Protein Binding; D055503:Protein Multimerization / Q000502:physiology*; D020836:Protein Structure, Quaternary; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051381:Rats; D051028:bcl-2-Associated X Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
C494826:BAX protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein