Structural insights into DNA sequence recognition by Type ISP restriction-modification enzymes.
Structural insights into DNA sequence recognition by Type ISP restriction-modification enzymes.
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DOI：
10.1093/nar/gkw154
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发表时间：
2016-05-19
影响因子：
14.9
通讯作者：
Saikrishnan K
中科院分区：
文献类型：
--
作者：
Kulkarni M;Nirwan N;van Aelst K;Szczelkun MD;Saikrishnan K
设计具有新序列特异性的限制性内切酶一直是一个尚未完成的挑战，可能是因为目标识别的复杂性。在这里，我们报告了ISP型限制性内切酶对靶标识别的详细分析。我们确定了与其靶标结合的ISP型酶LlaGI的结构，并将其与先前报道的与不同靶标LlaBIII结合的紧密同源物的结构进行了比较。比较表明，尽管这两种酶使用几乎相似的一组结构元素来识别靶标，但阅读碱基的残基不同。特异性的改变不仅是因为适当地取代了接触碱基的氨基酸，而且还因为位置不同的残基直接或通过水桥进行了新的接触。对552型isp酶的序列分析表明，LlaGI和LlaBIII参与靶标识别的结构元件在结构上是保守的，但在序列上却不那么保守。此外，这些结构元件中的残基位置受到强烈的进化约束，突显了这些区域的功能重要性。这项比较研究帮助破译了ISP型酶识别靶标的部分共识密码。
Engineering restriction enzymes with new sequence specificity has been an unaccomplished challenge, presumably because of the complexity of target recognition. Here we report detailed analyses of target recognition by Type ISP restriction-modification enzymes. We determined the structure of the Type ISP enzyme LlaGI bound to its target and compared it with the previously reported structure of a close homologue that binds to a distinct target, LlaBIII. The comparison revealed that, although the two enzymes use almost a similar set of structural elements for target recognition, the residues that read the bases vary. Change in specificity resulted not only from appropriate substitution of amino acids that contacted the bases but also from new contacts made by positionally distinct residues directly or through a water bridge. Sequence analyses of 552 Type ISP enzymes showed that the structural elements involved in target recognition of LlaGI and LlaBIII were structurally well-conserved but sequentially less-conserved. In addition, the residue positions within these structural elements were under strong evolutionary constraint, highlighting the functional importance of these regions. The comparative study helped decipher a partial consensus code for target recognition by Type ISP enzymes.