Crystal structure of Cas1 from Archaeoglobus fulgidus and characterization of its nucleolytic activity
Crystal structure of Cas1 from Archaeoglobus fulgidus and characterization of its nucleolytic activity
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DOI：
10.1016/j.bbrc.2013.10.122
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发表时间：
2013-11-29
影响因子：
3.1
通讯作者：
Kim, Jeong-Sun
中科院分区：
文献类型：
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作者：
Kim, Tae-Yang;Shin, Minsang;Kim, Jeong-Sun
CRISPR和CRISPR相关蛋白（Cas）参与细菌对有害遗传物质的获得性免疫。Cas1在含有CRISPR的微生物中普遍存在，并且应该识别和切割外源核酸，并使用尚未鉴定的机制将切割的片段整合到宿主基因组中。然而，所有已报道的Cas1s都没有显示出溶核活性，这使得其作用仍然不清楚。荧光古生菌Cas1（AfCas1）的晶体结构显示出蝴蝶状的二聚体结构。在其他Cas1中的Glu、His和Asp的三个确认的溶核残基中的Asp被AfCas1中的Glu取代。此外，五个残基插入到两个环中的一个，这是接近催化中心的和无序的其他Cas1结构，部分覆盖的活性位点的AfCas1。尽管如此，体外试验表明，其核酸结合活性对测试的单链（ss）DNA，各种形式的双链（ds）DNA，或ssRNA与水解活性对ssRNA和dsDNA在金属离子依赖性的方式不受损。这些结果支持了所提出的Cas1在这种细菌免疫系统的早期阶段的功能。(C)2013 Elsevier Inc. All rights reserved.
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) and CRISPR-associated (Cas) proteins are involved in bacterial acquired immunity against incoming hazardous genetic materials. Cas1 is ubiquitous in CRISPR-containing microorganisms and supposed to recognize and cleave a foreign nucleic acid, and integrate the cleaved fragment into host genome using a yet unidentified mechanism. However, all the reported Cas1s did not show the nucleolytic activity, which makes its role still obscure. The elucidated crystal structure of Cas1 from Archaeoglobus fulgidus (AfCas1) shows a butterfly-like dimeric structure. The Asp out of three confirmed nucleolytic residues of Glu, His, and Asp in other Cas1s is replaced with Glu in AfCas1. Further, insertion of five residues into one of two loops, which are close to the catalytic center of and disordered in other Cas1 structures, partially covers the active site of AfCas1. Nonetheless, in vitro assays show that its nucleic acid-binding activity was not impaired against the tested single-stranded (ss) DNA, various forms of double-stranded (ds) DNA, or ssRNA with a hydrolyzing activity against ssRNA and dsDNA in a metal ion-dependent way. These results support the proposed Cas1's function at the early step of this bacterial immune system. (C) 2013 Elsevier Inc. All rights reserved.