GAL4 TRANSCRIPTION FACTOR IS NOT A ZINC FINGER BUT FORMS A ZN(II)2CYS6 BINUCLEAR CLUSTER
GAL4 TRANSCRIPTION FACTOR IS NOT A ZINC FINGER BUT FORMS A ZN(II)2CYS6 BINUCLEAR CLUSTER
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DOI：
10.1073/pnas.87.6.2077
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发表时间：
1990-03-01
影响因子：
11.1
通讯作者：
COLEMAN, JE
中科院分区：
文献类型：
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作者：
TAO, P;COLEMAN, JE
转录因子GAL 4的DNA结合结构域由62个N-末端残基组成并表示为GAL 4（62*），含有Cys-Xaa 2-Cys-Xaa 6-Cys-Xaa 6-Cys-Xaa 2-Cys-Xaa 6-Cys基序，其先前已显示结合两个Zn（II）或Cd（II）离子。Zn（II）或Cd（II）的结合对于GAL 4识别特定回文DNA序列是必需的，GAL 4结合半乳糖代谢酶基因上游的UASG序列。基于两个结合的113 Cd（II）离子的113 Cd NMR化学位移，我们提出了一个双核簇模型的Zn（II）结合子域。112 Cd（II）-和113 Cd（II）-取代的GAL 4（62*）衍生物的1H-113 Cd杂原子多量子NMR光谱和相敏双量子滤波1H相关光谱提供了直接证据，表明两个结合的113 Cd（II）离子仅由六个半胱氨酸残基配位，其中两个半胱氨酸残基形成两个113 Cd（II）离子之间的桥接配体。后者可从~ 1H-~（113）Cd J偶联模式中鉴别。因此，双核金属离子簇，而不是“锌指”是由GAL 4 DNA结合结构域的六个半胱氨酸残基形成的。该模型可以直接应用于其他八种真菌转录因子，这些转录因子已被证明含有类似间隔的Cys 6簇。apo-GAL 4（62*）的1H NMR光谱表明在除去Zn（II）或Cd（II）后金属结合亚结构域的构象波动。Cd（II）2-和Zn（II）2-含物种的GAL 4可以形成，和类似的1H NMR光谱表明类似的构象。
The DNA-binding domain of the transcription factor GAL4, consisting of the 62 N-terminal residues and denoted GAL4(62*), contains a Cys-Xaa2-Cys-Xaa6-Cys-Xaa6-Cys-Xaa2-Cys-Xaa6-Cys motif, which has been shown previously to bind two Zn(II) or Cd(II) ions. Binding of Zn(II) or Cd(II) is essential for the recognition by GAL4 of the specific palindromic DNA sequence to which it binds upstream of genes for galactose-metabolizing enzymes, the UASG sequence. On the basis of the 113Cd NMR chemical shifts of the two bound 113Cd(II) ions, we propose a binuclear cluster model for this Zn(II)-binding subdomain. 1H-113Cd heteronuclear multiple-quantum NMR spectroscopy and phase-sensitive double-quantum filtered 1H correlation spectroscopy of the 112Cd(II)- and 113Cd(II)-substituted GAL4(62*) derivatives provide direct evidence that the two bound 113Cd(II) ions are coordinated only by the six cysteine residues, two of which form bridging ligands between the two 113Cd(II) ions. The latter can be identified from the pattern of 1H-113Cd J coupling. Thus a binuclear metal ion cluster rather than a "zinc finger" is formed by the six cysteine residues of the GAL4 DNA-binding domain. This model can be directly applied to eight other fungal transcription factors which have been shown to contain similarly spaced Cys6 clusters. 1H NMR spectra of apo-GAL4(62*) suggest conformational fluctuation of the metal-binding subdomain upon removal of Zn(II) or Cd(II). Both Cd(II)2- and Zn(II)2-containing species of GAL4 can be formed, and the similar 1H NMR spectra suggest similar conformations.