Identification, Characterization, and Structure Analysis of the Cyclic di-AMP-binding PII-like Signal Transduction Protein DarA
Identification, Characterization, and Structure Analysis of the Cyclic di-AMP-binding PII-like Signal Transduction Protein DarA
复制标题
DOI：
10.1074/jbc.m114.619619
复制
发表时间：
2015-01-30
影响因子：
4.8
通讯作者：
Ficner, Ralf
中科院分区：
文献类型：
--
作者：
Gundlach, Jan;Dickmanns, Achim;Ficner, Ralf
背景：环二磷酸腺苷是真细菌中一种重要的第二信使。结果：在B中鉴定出c-di-AMP受体达拉A。枯草芽孢杆菌晶体结构和ITC数据揭示了达拉A的核苷酸特异性。结论：达拉A是一种与c-di-AMP结合后发生构象变化的P-II样蛋白。重要性：一种新的P-II样蛋白参与c-di-AMP信号转导，环二聚体AMP核苷酸c-di-AMP是枯草芽孢杆菌中必需的第二信使。我们已经鉴定出蛋白质达拉A是B中主要的c-di-AMP受体之一。枯草芽孢杆菌晶体结构分析表明，达拉A与P-II信号转导蛋白高度同源。与P-II蛋白相反，功能上重要的B环和T环的大小互换。达拉A是一种同源三聚体，它结合三个c-di-AMP分子，每个分子位于两个亚基之间的口袋中。我们证明了达拉A能够结合c-di-AMP和低亲和力的环GMP-AMP（33-cGAMP），但不能结合c-di-GMP或23-cGAMP。一致的晶体结构表明，在配体结合口袋中，只有一个腺嘌呤是高度特异性识别，而其他腺嘌呤的口袋似乎是混杂的。与同源无配体的达拉A结构的比较表明，c-di-AMP结合伴随着达拉A中B环的折叠和位置的构象变化。
Background: Cyclic di-AMP is an essential second messenger in eubacteria. Results: The c-di-AMP receptor DarA was identified in B. subtilis. The crystal structure and ITC data revealed the nucleotide specificity of DarA. Conclusion: DarA is a P-II-like protein that undergoes conformational changes upon c-di-AMP binding. Significance: A novel P-II-like protein is involved in c-di-AMP signaling.The cyclic dimeric AMP nucleotide c-di-AMP is an essential second messenger in Bacillus subtilis. We have identified the protein DarA as one of the prominent c-di-AMP receptors in B. subtilis. Crystal structure analysis shows that DarA is highly homologous to P-II signal transducer proteins. In contrast to P-II proteins, the functionally important B- and T-loops are swapped with respect to their size. DarA is a homotrimer that binds three molecules of c-di-AMP, each in a pocket located between two subunits. We demonstrate that DarA is capable to bind c-di-AMP and with lower affinity cyclic GMP-AMP (33-cGAMP) but not c-di-GMP or 23-cGAMP. Consistently the crystal structure shows that within the ligand-binding pocket only one adenine is highly specifically recognized, whereas the pocket for the other adenine appears to be promiscuous. Comparison with a homologous ligand-free DarA structure reveals that c-di-AMP binding is accompanied by conformational changes of both the fold and the position of the B-loop in DarA.