Crystal structure and nucleic acid-binding activity of the CRISPR-associated protein Csx1 of Pyrococcus furiosus
Crystal structure and nucleic acid-binding activity of the CRISPR-associated protein Csx1 of Pyrococcus furiosus
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DOI：
10.1002/prot.24183
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发表时间：
2013-02-01
影响因子：
2.9
通讯作者：
Oh, Byung-Ha
中科院分区：
文献类型：
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作者：
Kim, Young Kwan;Kim, Yeon-Gil;Oh, Byung-Ha
在许多原核生物中，被称为成簇规则间隔短回文重复序列（CRISPR）和CRISPR相关（CAS）基因的染色体基因座包含针对入侵的质粒和质粒的获得性免疫防御系统。虽然已经鉴定了许多不同的Cas蛋白家族，但其大多数组分的确切生化功能仍有待确定。在这项研究中，我们报告了PF 1127的晶体结构，PF 1127是一种由480个氨基酸组成的Cas蛋白，属于Csx 1家族。PF 1127的C-末端结构域具有独特的β-发夹结构，其突出于α-螺旋之外并含有几个带正电荷的残基。我们证明PF 1127结合双链DNA和RNA，这种活性需要一个完整的β-发夹，并涉及蛋白质的同源二聚化。相比之下，来自硫磺硫化叶菌P2的另一种Csx 1蛋白由377个氨基酸组成，不具有β-发夹结构，并且在相同的实验条件下不表现出DNA结合特性。值得注意的是，这两个Csx 1蛋白的C-末端结构域是非常多样化的，在保守的N-末端结构域，这似乎发挥了共同的作用，在同源二聚化的蛋白质。因此，虽然P. furiosus Csx 1被鉴定为核酸结合蛋白，但预测其他Csx 1蛋白表现出不同的个体生物化学活性。Proteins 2013. (C)2012 Wiley Periodicals，Inc.
In many prokaryotic organisms, chromosomal loci known as clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (CAS) genes comprise an acquired immune defense system against invading phages and plasmids. Although many different Cas protein families have been identified, the exact biochemical functions of most of their constituents remain to be determined. In this study, we report the crystal structure of PF1127, a Cas protein of Pyrococcus furiosus DSM 3638 that is composed of 480 amino acids and belongs to the Csx1 family. The C-terminal domain of PF1127 has a unique beta-hairpin structure that protrudes out of an a-helix and contains several positively charged residues. We demonstrate that PF1127 binds double-stranded DNA and RNA and that this activity requires an intact beta-hairpin and involve the homodimerization of the protein. In contrast, another Csx1 protein from Sulfolobus solfataricus P2 that is composed of 377 amino acids does not have the beta-hairpin structure and exhibits no DNA-binding properties under the same experimental conditions. Notably, the C-terminal domain of these two Csx1 proteins is greatly diversified, in contrast to the conserved N-terminal domain, which appears to play a common role in the homodimerization of the protein. Thus, although P. furiosus Csx1 is identified as a nucleic acid-binding protein, other Csx1 proteins are predicted to exhibit different individual biochemical activities. Proteins 2013. (C) 2012 Wiley Periodicals, Inc.