Exploring the DNA-recognition potential of homeodomains
Exploring the DNA-recognition potential of homeodomains
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DOI：
10.1101/gr.139014.112
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发表时间：
2012-10-01
期刊：
影响因子：
7
通讯作者：
Wolfe, Scot A.
中科院分区：
文献类型：
--
作者：
Chu, Stephanie W.;Noyes, Marcus B.;Wolfe, Scot A.
大多数DNA结合结构域（DBD）家族的识别潜力仍然相对未开发。同源结构域（HD），像许多其他家庭的DBD，显示有限的多样性，在他们的首选识别序列。为了探索HD的识别潜力，我们利用细菌选择系统从随机化文库中分离HD变体，所述变体与64个可能的3'三联体位点中的每一个（即，TAANNN）。这些选择中的大多数产生了在特定识别位置具有过度代表的残基的HD组，这意味着选择了特定的结合剂。随后表征了151种代表性HD变体的DNA结合特异性，鉴定了优先识别这些靶位点中的44个的HD。这些变体中的许多含有在现存的HD中不常见或不存在的特异性决定簇的新组合。这些新的决定簇，当移植到不同的HD骨架，产生相应的特异性改变。这些信息被用来创建更明确的HD识别模型，这可以告知预测的转录调控网络现存的HD或工程的HD与新的DNA识别潜力。恢复HD识别序列的多样性提出了重要的问题，健身障碍，限制了自然系统中的替代识别方式的演变。
The recognition potential of most families of DNA-binding domains (DBDs) remains relatively unexplored. Homeodomains (HDs), like many other families of DBDs, display limited diversity in their preferred recognition sequences. To explore the recognition potential of HDs, we utilized a bacterial selection system to isolate HD variants, from a randomized library, that are compatible with each of the 64 possible 3' triplet sites (i.e., TAANNN). The majority of these selections yielded sets of HDs with overrepresented residues at specific recognition positions, implying the selection of specific binders. The DNA-binding specificity of 151 representative HD variants was subsequently characterized, identifying HDs that preferentially recognize 44 of these target sites. Many of these variants contain novel combinations of specificity determinants that are uncommon or absent in extant HDs. These novel determinants, when grafted into different HD backbones, produce a corresponding alteration in specificity. This information was used to create more explicit HD recognition models, which can inform the prediction of transcriptional regulatory networks for extant HDs or the engineering of HDs with novel DNA-recognition potential. The diversity of recovered HD recognition sequences raises important questions about the fitness barrier that restricts the evolution of alternate recognition modalities in natural systems.