Functional characterization of iron-substituted neural zinc finger factor 1: metal and DNA binding
Functional characterization of iron-substituted neural zinc finger factor 1: metal and DNA binding
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DOI：
10.1007/s00775-010-0626-1
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发表时间：
2010-05-01
影响因子：
3
通讯作者：
Cymet, Holly J.
中科院分区：
文献类型：
--
作者：
Besold, Angelique N.;Lee, Seung Jae;Cymet, Holly J.
神经锌指因子1 （Neural zinc finger factor 1, NZF-1）是一种参与神经元发育的非经典锌指蛋白。NZF-1含有一个独特的CCHHC锌结合域的多个拷贝，该域识别β -视黄酸受体基因中的启动子元件，称为β -视黄酸受体元件（β - rare）。先前的研究已经确定，与锌结合的NZF-1的双结构域片段足以进行特定的DNA结合。神经系统的正常功能在很大程度上依赖于铁，这种高度氧化还原活性金属的调节不当会造成严重后果。有几类锌指蛋白被证明可以结合其他金属离子，包括铁离子。为了确定亚铁是否可以与NZF-1的金属结合位点协调并评估这种协调的功能后果，我们制备了一个包含两个锌结合结构域的NZF-1片段，即NZF-1双指（NZF-1- df）。紫外-可见光谱实验表明，Fe（II）能够与NZF-1-DF结合。在与铁（II）或锌（II）重组后，NZF-1-DF选择性地与靶β - rare DNA序列紧密结合（纳米摩尔亲和力），而载脂蛋白NZF-1-DF不与DNA结合，而是聚集。
Neural zinc finger factor 1 (NZF-1) is a nonclassical zinc finger protein involved in neuronal development. NZF-1 contains multiple copies of a unique CCHHC zinc-binding domain that recognize a promoter element in the beta-retinoic acid receptor gene termed beta-retinoic acid receptor element (beta-RARE). Previous studies have established that a two-domain fragment of NZF-1 bound with zinc is sufficient for specific DNA binding. Proper functioning of the nervous system relies heavily on iron and misregulation of this highly redox active metal has serious consequences. Several classes of zinc finger proteins have been shown to bind other metal ions, including iron. To determine if ferrous iron can coordinate to the metal-binding sites of NZF-1 and assess the functional consequences of such coordination, a fragment of NZF-1 that contains two zinc-binding domains, NZF-1 double finger (NZF-1-DF), was prepared. UV-vis spectroscopy experiments demonstrated that Fe(II) is capable of binding to NZF-1-DF. Upon reconstitution with either Fe(II) or Zn(II), NZF-1-DF binds selectively and tightly (nanomolar affinity) to its target beta-RARE DNA sequence, whereas apo-NZF-1-DF does not bind to DNA and instead aggregates.