期刊
NUCLEIC ACIDS RESEARCH
卷 45, 期 11, 页码 6960-6970出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkx342
关键词
-
资金
- National Institutes of Health [GM9886, GM105691]
Transcription activator-like effectors (TALEs) recognize their DNA targets via tandem repeats, each specifying a single nucleotide base in a one-to-one sequential arrangement. Due to this modularity and their ability to bind long DNA sequences with high specificity, TALEs have been used in many applications. Contributions of individual repeat-nucleotide associations to affinity and specificity have been characterized. Here, using in vitro binding assays, we examined the relationship between the number of repeats in a TALE and its affinity, for both target and non-target DNA. Each additional repeat provides extra binding energy for the target DNA, with the gain decaying exponentially such that binding energy saturates. Affinity for non-target DNA also increases non-linearly with the number of repeats, but with a slower decay of gain. The difference between the effect of length on affinity for target versus nontarget DNA manifests in specificity increasing then diminishing with increasing TALE length, peaking between 15 and 19 repeats. Modeling across different hypothetical saturation levels and rates of gain decay, reflecting different repeat compositions, yielded a similar range of specificity optima. This range encompasses the mean and median length of native TALEs, suggesting that these proteins as a group have evolved for maximum specificity.
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