Zha H, Aimé-Sempé C, Sato T, Reed J C
La Jolla Cancer Research Foundation, California 92037, USA.
J Biol Chem. 1996 Mar 29;271(13):7440-4. doi: 10.1074/jbc.271.13.7440.
Most members of the Bcl-2 protein family of apoptosis regulating proteins contain two evolutionarily conserved domains, termed BH1 and BH2. Both BH1 and BH2 in the Bcl-2 protein are required for its function as an inhibitor of cell death and for heterodimerization with the proapoptotic protein Bax. In this report, we mapped the region in Bax required for heterodimerization with Bcl-2 and homodimerization with Bax, using yeast two-hybrid and in vitro protein-protein interaction assays. Neither the BH1 nor the BH2 domain of Bax was required for binding to the wild-type Bcl-2 and Bax proteins. Moreover, Bax (deltaBH1) and Bax (deltaBH2) mutant proteins bound efficiently to themselves and each other, further confirming the lack of requirement for BH1 and BH2 for Bax/Bax homodimerization. Bax/Bax homodimerization was not dependent on the inclusion of the NH2-terminal 58 amino acids of the Bax protein in each dimerization partner, unlike Bcl-2/Bcl-2 homodimers which involve head-to-tail interactions between the region of Bcl-2 where BH1 and BH2 resides, and an NH2-terminal domain in Bcl-2 that contains another domain BH4 which is conserved among antiapoptotic members of the Bcl-2 family. Similarly, heterodimerization with Bcl-2 occurred without the NH2-terminal domain of either Bax or Bcl-2, suggesting a tail-to-tail interaction. The essential region in Bax required for both homodimerization with Bax and heterodimerization with Bcl-2 was mapped to residues 59-101. This region in Bax contains a stretch of 15 amino acids that is highly homologous in several members of the Bcl-2 protein family, suggesting the existence of a novel functional domain which we have termed BH3. Deletion of this 15-amino acid region abolished the ability of Bax to dimerize with itself and to heterodimerize with Bcl-2. The findings suggest that the structural features of Bax and Bcl-2 that allow them to participate in homo-and heterodimerization phenomena are markedly different, despite their amino-acid sequence similarity.
凋亡调节蛋白的Bcl-2蛋白家族的大多数成员包含两个进化上保守的结构域，称为BH1和BH2。Bcl-2蛋白中的BH1和BH2对于其作为细胞死亡抑制剂的功能以及与促凋亡蛋白Bax的异源二聚化都是必需的。在本报告中，我们使用酵母双杂交和体外蛋白质-蛋白质相互作用分析，确定了Bax中与Bcl-2异源二聚化以及与Bax同源二聚化所需的区域。Bax的BH1和BH2结构域都不是与野生型Bcl-2和Bax蛋白结合所必需的。此外，Bax（deltaBH1）和Bax（deltaBH2）突变蛋白能有效地自身结合并相互结合，进一步证实了Bax/Bax同源二聚化不需要BH1和BH2。与Bcl-2/Bcl-2同源二聚体不同，Bax/Bax同源二聚化不依赖于每个二聚化伙伴中包含Bax蛋白的NH2末端58个氨基酸，Bcl-2/Bcl-2同源二聚体涉及BH1和BH2所在的Bcl-2区域与Bcl-2中包含另一个在Bcl-2家族抗凋亡成员中保守的BH4结构域的NH2末端结构域之间的头对头相互作用。同样，Bax与Bcl-2的异源二聚化在没有Bax或Bcl-2的NH2末端结构域的情况下也会发生，这表明是尾对尾相互作用。Bax中与Bax同源二聚化以及与Bcl-2异源二聚化所需的关键区域被定位到第59 - 101位氨基酸残基。Bax中的这个区域包含一段15个氨基酸的序列，在Bcl-2蛋白家族的几个成员中高度同源，这表明存在一个我们称为BH3的新功能结构域。删除这15个氨基酸区域消除了Bax自身二聚化以及与Bcl-2异源二聚化的能力。这些发现表明，尽管Bax和Bcl-2的氨基酸序列相似，但它们参与同源和异源二聚化现象的结构特征明显不同。
