Structural and functional characterizations reveal the importance of a zinc binding domain in Bloom's syndrome helicase.
Structural and functional characterizations reveal the importance of a zinc binding domain in Bloom's syndrome helicase.
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DOI：
10.1093/nar/gki619
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发表时间：
2005
影响因子：
14.9
通讯作者：
Xi XG
中科院分区：
文献类型：
--
作者：
Guo RB;Rigolet P;Zargarian L;Fermandjian S;Xi XG
布卢姆综合征（BS）是一种常染色体隐性遗传的人类疾病，其特征是基因组不稳定和易患多种癌症。在BS中突变的基因BLM编码含有三个结构域的蛋白质：N-末端结构域，其功能仍然难以捉摸，解旋酶结构域，其特征在于在广泛的解旋酶中保守的七个“签名”基序和C-末端延伸，其可以进一步分为两个亚结构域：RecQ-Ct和HRDC。RecQ-Ct结构域似乎是必不可少的，因为在BS患者中发现了改变该结构域内高度保守的半胱氨酸残基的两个点突变。我们在此报告，BLM含有锌离子。建模研究表明，RecQ-Ct结构域内的四个保守的半胱氨酸残基协调该锌离子，随后的诱变研究进一步证实了这一预测。生化和生物物理研究表明，ATP酶，解旋酶和DNA结合活性的突变体的严重修改。野生型和突变体蛋白的结构分析表明，半胱氨酸残基的改变不会显着改变整体构象。ATP酶和解旋酶活性的观察到的缺陷被推断为DNA结合的折衷结果。我们的研究结果暗示了这个锌结合域在DNA结合和蛋白质构象中的重要作用。它们可能是理解BS疾病分子基础的关键。
Bloom's syndrome (BS) is an autosomal recessive human disorder characterized by genomic instability and a predisposition to a wide variety of cancers. The gene mutated in BS, BLM, encodes a protein containing three domains: an N-terminal domain whose function remains elusive, a helicase domain characterized by seven ‘signature’ motifs conserved in a wide range of helicases and a C-terminal extension that can be further divided into two sub-domains: RecQ-Ct and HRDC. The RecQ-Ct domain appears essential because two point-mutations altering highly conserved cysteine residues within this domain have been found in BS patients. We report herein that BLM contains a zinc ion. Modelling studies suggest that four conserved cysteine residues within the RecQ-Ct domain coordinate this zinc ion and subsequent mutagenesis studies further confirm this prediction. Biochemical and biophysical studies show that the ATPase, helicase and DNA binding activities of the mutants are severely modified. Structural analysis of both wild-type and mutant proteins reveal that alteration of cysteine residues does not significantly change the overall conformation. The observed defects in ATPase and helicase activities were inferred to result from a compromise of DNA binding. Our results implicate an important role of this zinc binding domain in both DNA binding and protein conformation. They could be pivotal for understanding the molecular basis of BS disease.
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