Toroidal Structure and DNA Cleavage by the CRISPR-Associated [4Fe-4S] Cluster Containing Cas4 Nuclease SSO0001 from Sulfolobus solfataricus

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• 作者：Sofia Lemak ; Natalia Beloglazova ; Boguslaw Nocek ; Tatiana Skarina ; Robert Flick ; Greg Brown ; Ana Popovic ; Andrzej Joachimiak ; Alexei Savchenko ; Alexander F. Yakunin
• 刊名：Journal of the American Chemical Society
• 出版年：2013
• 出版时间：November 20, 2013
• 年：2013
• 卷：135
• 期：46
• 页码：17476-17487
• 全文大小：801K
• 年卷期：v.135,no.46(November 20, 2013)
• ISSN：1520-5126

文摘

Cas4 proteins, a core protein family associated with the microbial system of adaptive immunity CRISPR, are predicted to function in the adaptation step of the CRISPR mechanism. Here we show that the Cas4 protein SSO0001 from the archaeon Sulfolobus solfataricus has metal-dependent endonuclease and 5鈥测啋3鈥?exonuclease activities against single-stranded DNA, as well as ATP-independent DNA unwinding activity toward double-stranded DNA. The crystal structure of SSO0001 revealed a decameric toroid formed by five dimers with each protomer containing one [4Fe-4S] cluster and one Mn²⁺ ion bound in the active site located inside the internal tunnel. The conserved RecB motif and four Cys residues are important for DNA binding and cleavage activities, whereas DNA unwinding depends on several residues located near the [4Fe-4S] cluster. Our results suggest that Cas4 proteins might contribute to the addition of novel CRISPR spacers through the formation of 3鈥?DNA overhangs and to the degradation of foreign DNA.