Chi Haotian, McMahon Stephen, Daniel-Pedersen Lukas, Graham Shirley, Gloster Tracey M, White Malcolm F
School of Biology, University of St Andrews, St Andrews, Fife KY16 9ST, United Kingdom.
Nucleic Acids Res. 2025 Jul 8;53(13). doi: 10.1093/nar/gkaf655.
Type III CRISPR systems detect non-self RNA and activate the enzymatic Cas10 subunit, which generates nucleotide second messengers for activation of ancillary effectors. Although most signal via cyclic oligoadenylate, an alternative class of signalling molecule SAM-AMP, formed by conjugating ATP and S-adenosylmethionine, was described recently. SAM-AMP activates a trans-membrane effector of the CorA magnesium transporter family to provide anti-phage defence. Intriguingly, immunity also requires SAM-AMP degradation by means of a specialized CRISPR-encoded NrN family phosphodiesterase in Bacteroides fragilis. In Clostridium botulinum, the nrn gene is replaced by a gene encoding a SAM-AMP lyase. Here, we investigate the structure and activity of C. botulinum SAM-AMP lyase, which can substitute for the nrn gene to provide CorA-mediated immunity in Escherichia coli. The structure of SAM-AMP lyase bound to its reaction product 5'-methylthioadenosine-AMP reveals key details of substrate binding and turnover by this PII superfamily protein. Bioinformatic analysis revealed a phage-encoded SAM-AMP lyase that degrades SAM-AMP efficiently in vitro, consistent with an anti-CRISPR function.
III型CRISPR系统可检测非自身RNA并激活酶促Cas10亚基，该亚基会生成核苷酸第二信使以激活辅助效应蛋白。尽管大多数信号是通过环状寡腺苷酸传递的，但最近发现了另一类由ATP与S-腺苷甲硫氨酸结合形成的信号分子SAM-AMP。SAM-AMP激活了CorA镁转运蛋白家族的一种跨膜效应蛋白，以提供抗噬菌体防御。有趣的是，在脆弱拟杆菌中，免疫作用还需要通过一种专门的CRISPR编码的NrN家族磷酸二酯酶来降解SAM-AMP。在肉毒梭菌中，nrn基因被一个编码SAM-AMP裂解酶的基因所取代。在此，我们研究了肉毒梭菌SAM-AMP裂解酶的结构和活性，该酶可替代nrn基因，在大肠杆菌中提供CorA介导的免疫作用。与反应产物5'-甲硫基腺苷-AMP结合的SAM-AMP裂解酶的结构揭示了这种PII超家族蛋白底物结合和周转的关键细节。生物信息学分析揭示了一种噬菌体编码的SAM-AMP裂解酶，它在体外能有效降解SAM-AMP，这与一种抗CRISPR功能一致。
