Mutation to Bax beyond the BH3 Domain Disrupts Interactions with Pro-survival Proteins and Promotes Apoptosis
Mutation to Bax beyond the BH3 Domain Disrupts Interactions with Pro-survival Proteins and Promotes Apoptosis
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DOI：
10.1074/jbc.m110.161281
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发表时间：
2011-03-04
影响因子：
4.8
通讯作者：
Colman, Peter M.
中科院分区：
文献类型：
--
作者：
Czabotar, Peter E.;Lee, Erinna F.;Colman, Peter M.
Bcl2家族中的促生存成员通过直接与Bax的相互作用或间接地通过隔离激活BH3的蛋白，或两者兼而有之，抑制Bax的促凋亡活性。促进细胞凋亡的Bax突变可以提供对Bax是如何调控的洞察。在这里，我们描述了促生存蛋白Mcl-1和Bclx(L)与BAX中包含BH3结构域的34肽的复合体的晶体结构。这些结构揭示了来自BaxBH3结构域的四个标志性疏水氨基酸与有利于生存的蛋白质的BH3结合槽之间的典型相互作用。在这两种结构中，Bax多肽的Met-74与BH3结合槽以第五疏水相互作用的方式与BH3结合。不同的Bax Met-74突变体破坏了Bax和所有促生存蛋白之间的相互作用，但这些Bax突变体保持了促凋亡活性。用几个Bax Met-74突变体重组的Bax/Bak缺陷的小鼠胚胎成纤维细胞对BH3模拟化合物ABT-737比表达野生型Bax的细胞更敏感。此外，即使在没有外部凋亡刺激的情况下，表达Bax Met-74突变体的细胞在集落分析中的存活率也较低。这些结果支持这样一种模型，即促生存的Bcl-2蛋白直接抑制Bax是细胞凋亡的屏障。
Pro-survival members of the Bcl-2 family of proteins restrain the pro-apoptotic activity of Bax, either directly through interactions with Bax or indirectly by sequestration of activator BH3-only proteins, or both. Mutations in Bax that promote apoptosis can provide insight into how Bax is regulated. Here, we describe crystal structures of the pro-survival proteins Mcl-1 and Bcl-x(L) in complex with a 34-mer peptide from Bax that encompasses its BH3 domain. These structures reveal canonical interactions between four signature hydrophobic amino acids from the BaxBH3 domain and the BH3-binding groove of the pro-survival proteins. In both structures, Met-74 from the Bax peptide engages with the BH3-binding groove in a fifth hydrophobic interaction. Various Bax Met-74 mutants disrupt interactions between Bax and all pro-survival proteins, but these Bax mutants retain pro-apoptotic activity. Bax/Bak-deficient mouse embryonic fibroblast cells reconstituted with several Bax Met-74 mutants are more sensitive to the BH3 mimetic compound ABT-737 as compared with cells expressing wild-type Bax. Furthermore, the cells expressing Bax Met-74 mutants are less viable in colony assays even in the absence of an external apoptotic stimulus. These results support a model in which direct restraint of Bax by pro-survival Bcl-2 proteins is a barrier to apoptosis.