Structural Insights into the Distinct Binding Mode of Cyclic Di-AMP with SaCpaA_RCK

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• 作者：Ko-Hsin Chin ; Juin-Ming Liang ; Jauo-Guey Yang ; Min-Shao Shih ; Zhi-Le Tu ; Yu-Chuang Wang ; Xing-Han Sun ; Nien-Jen Hu ; Zhao-Xun Liang ; J. Maxwell Dow ; Robert P. Ryan ; Shan-Ho Chou
• 刊名：Biochemistry
• 出版年：2015
• 出版时间：August 11, 2015
• 年：2015
• 卷：54
• 期：31
• 页码：4936-4951
• 全文大小：1070K
• ISSN：1520-4995

文摘

Cyclic di-AMP (c-di-AMP) is a relatively new member of the family of bacterial cyclic dinucleotide second messengers. It has attracted significant attention in recent years because of the abundant roles it plays in a variety of Gram-positive bacteria. The structural features that allow diverse bacterial proteins to bind c-di-AMP are not fully understood. Here we report the biophysical and structural studies of c-di-AMP in complex with a bacterial cation鈥損roton antiporter (CpaA) RCK (regulator of the conductance of K⁺) protein from Staphylococcus aureus (Sa). The crystal structure of the SaCpaA_RCK C-terminal domain (CTD) in complex with c-di-AMP was determined to a resolution of 1.81 脜. This structure revealed two well-liganded water molecules, each interacting with one of the adenine bases by a unique H₂O_lp鈥撓€ interaction to stabilize the complex. Sequence blasting using the SaCpaA_RCK primary sequence against the bacterial genome database returned many CpaA analogues, and alignment of these sequences revealed that the active site residues are all well-conserved, indicating a universal c-di-AMP binding mode for CpaA_RCK. A proteoliposome activity assay using the full-length SaCpaA membrane protein indicated that c-di-AMP binding alters its antiporter activity by approximately 40%. A comparison of this structure to all other reported c-di-AMP鈥搑eceptor complex structures revealed that c-di-AMP binds to receptors in either a 鈥淯-shape鈥?or 鈥淰-shape鈥?mode. The two adenine rings are stabilized in the inner interaction zone by a variety of CH鈭捪€, cation鈭捪€, backbone鈭捪€, or H₂O_lp鈥撓€ interaction, but more commonly in the outer interaction zone by hydrophobic CH鈭捪€ or 蟺鈥撓€ interaction. The structures determined to date provide an understanding of the mechanisms by which a single c-di-AMP can interact with a variety of receptor proteins, and how c-di-AMP binds receptor proteins in a special way different from that of c-di-GMP.