序列母题
主题(音乐)
突变体
生物
转录因子
结合位点
共识序列
结构母题
遗传学
分子生物学
细胞生物学
肽序列
DNA
基因
生物化学
物理
声学
作者
Dongyun Hao,Kazuhiko Yamasaki,Akinori Sarai,Masaru Ohme‐Takagi
出处
期刊:Biochemistry
[American Chemical Society]
日期:2002-03-07
卷期号:41 (13): 4202-4208
被引量:109
摘要
Arabidopsis ERF proteins such as DREB1, DREB2, and CBF1 bind to the dehydration-responsive element (DRE), which has the sequence TACCGACAT. Mutation analyses reveal that a central 5 bp CCGAC core of the DRE is the minimal sequence motif (designated as the DRE motif in this paper), to which the ERF domain fragment of CBF1 (CBF1-F) binds specifically with a binding Kd at the nanomolar level. In contrast, the ERF domain fragment of the tobacco ERF2 (NtERF2-F) does not interact with the DRE motif, but restrictedly recognizes the sequence containing a minimal 6 bp GCCGCC motif (designated as the GCC motif in this paper). However, CBF1-F binds to the GCC motif with a binding activity similar to its binding activity for the DRE motif. These in vitro binding variations were further demonstrated through reporter cotransformation assays, suggesting that the DRE and GCC motifs are two similar sequence motifs sharing a common core region of CCGNC with a discriminating guanine base at the 5'-end of the GCC motif. Binding analyses with the mutated ERF domain show that such a unique binding of NtERF2-F to the GCC motif can be altered by the substitution of A14 with valine in β-strand 2 of its ERF domain, the mutant NtERF2-F, ERFav, acquiring a binding to the DRE motif with a Kd comparable to that for CBF1-F binding to the DRE motif. This demonstrates that A14 is an important determinant of the NtERF2-F binding specificity. A possible mechanism of the binding specificity determination is discussed.
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