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首页> 外文期刊>Journal of Molecular Biology >The solution structure of the Leu22 -> Val mutant AREA DNA binding domain complexed with a TGATAG core element defines a role for hydrophobic packing in the determination of specificity
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The solution structure of the Leu22 -> Val mutant AREA DNA binding domain complexed with a TGATAG core element defines a role for hydrophobic packing in the determination of specificity

机译：与TGATAG核心元件复合的Leu22-> Val突变体AREA DNA结合结构域的溶液结构定义了疏水性包装在确定特异性中的作用

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摘要

The seemingly innocuous leucine-to-valine mutation at Position 22 of the AREA DNA binding domain results in dramatic changes in the in vivo expression Profile of genes controlled by this GATA transcription factor. This is associated with a Preference of the Leu22 --> Val mutant for TGATAG sites over (A/C)GATAG sites. Quantitative gel retardation assays confirm this observation and show that the Leu22 --> Val mutant AREA DNA binding domain has a similar to 30-fold lower affinity than the wild-type domain for a 13 base-pair oligonucleotide containing the wild-type CGATAG target. To gain insight into the measured affinity data and further explore sequence specificity of the AREA protein, the solution structure of a complex between the Leu22 --> Val mutant AREA DNA binding domain and a 13 base-pair oligonucleotide containing its physiologically relevant TGATAG target sequence has been determined by multidimensional nuclear magnetic resonance spectroscopy. Comparison of this structure with that of the wild-type AREA DNA binding domain complexed to its cognate CGATAG target site shows how subtle changes in amino acid side-chain length and hydrophobic packing car, affect affinity and specificity GATA-containing sequences, and how changes in DNA sequence be compensated for by changes in protein sequence. (C) 1998 Academic Press Limited. [References: 28]
机译：在AREA DNA结合结构域第22位看似无害的亮氨酸-缬氨酸突变导致该GATA转录因子控制的基因的体内表达谱发生显着变化。与（A / C）GATAG位点相比，TGATAG位点更倾向于Leu22-> Val突变体。定量凝胶阻滞测定法证实了这一发现，并表明对于包含野生型CGATAG靶标的13个碱基对的寡核苷酸，Leu22-> Val突变型AREA DNA结合域的亲和力比野生型域低30倍。 。为了深入了解测得的亲和力数据并进一步探索AREA蛋白的序列特异性，Leu22-> Val突变型AREA DNA结合结构域与包含其生理相关TGATAG靶序列的13个碱基对寡核苷酸之间的复合物的溶液结构已经通过多维核磁共振波谱法确定。将该结构与其复合的CGATAG目标位点复合的野生型AREA DNA结合结构域的结构进行比较，可以看出氨基酸侧链长度和疏水性包装载体的细微变化如何影响含GATA的亲和力和特异性序列，以及如何变化DNA序列中的蛋白被蛋白质序列的变化所补偿。 （C）1998 Academic Press Limited。 [参考：28]

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