Research News
Studies from Dr. Feng Shao's laboratory identified new target of enteropathogenic bacteria effector protein.
On Nov. 20, 2014, Studies from Dr. Feng Shao's laboratory at National Institute of Biological Sciences (NIBS), Beijing, elucidate structural mechanisms for a novel cysteine methyltransferase effector NleE from enteropathogenic E. coli and identify DNA repair protein ZRANB3 as a novel methylation substrate. The work entitled "Structure and Specificity of the Bacterial Cysteine Methyltransferase Effector NleE Suggests a Novel Substrate in Human DNA Repair Pathway" is published online in the journal PLOS Pathogens.
In a previous study from Dr. Shao's laboratory (Nature 481: 204-208, 2012), they discover that NleE inhibits NF-kB signaling by methylating a specific Zn-coordinating cysteine in host TAB2/3 and disrupting their ubiquitin-chain binding activity. That study for the first time demonstrates that cysteine methylation regulates eukaryotic signal transduction. As NleE shows no sequence homology to all known methyltransferases, deep understanding of NleE structure and function is important for further study of this new posttranslational modification.
In the current study, researchers in the Shao group first determined the crystal structure of NleE in complex with the methyl donor SAM, which reveals a unique a+b Rossmann-like fold. The whole molecule bears a sandwich shape. The SAM is mainly contacted by three critical residues at the bottom of a pocket in NleE. Structure-based molecular docking and dynamic simulation analyses indicates that Cys673, among the four Zn-coordinating cysteine in TAB2, is spatially and energetically favorable for being methylated. The authors also identify an N-terminal sequence region in TAB3 that is required for efficient substrate recognition.
Given the wide prevalence of Zn-finger motifs in eukaryotes, the researchers further checked ~ 50 Zn-finger motifs and found that NleE could also methylate ZRANB3 on its Cys630. The modification efficiency is comparable to that of TAB2/3. ZRANB3 functions in the DNA repair pathway and is important for maintaining genome stability. Previous studies have indicated that ZRANB3 is recruited to DNA damage site, which may involve Zn-finger motif binding to ubiquitin chains on PCNA. The researchers also discover that NleE methylation abolishes the ubiquitin-chain binding activity of ZRANB3, raising a stimulating question whether EPEC infection may lead to host genome instability.
The study for the first time solved the structure of a pathogen-derived methyltransferase effector NleE, performed in-depth structural and functional comparison with known methyltransferases, and therefore elucidated the mechanism of NleE-catalyzed cysteine methylation. The study further identified a new substrate (ZRANB3) for NleE. Taking advantage of the unique activity of NleE, the authors were able to show that ZRANB3 binding to ubiquitin chains and recruitment to DNA damage sites are mutually independent processes. This highlights the emerging concept of using pathogenic effectors for dissecting host cellular processes.
Postdoc fellows Qing Yao and Li Zhang from the Shao group are co-first authors of this paper; Dr. Xiaobo Wan from the Niu Huang laboratory and rotation student Jing Chen have also made significant contributions; other contributors include Liyan Hu (the Shao group), Xiaojun Ding, Lin Li and Dr. She Chen at NIBS proteomics center, Jayashree Karar, Hongzhuang Peng and Dr. Frank J. Rauscher III at Wistar Institute(USA), as well as Dr. Niu Huang at NIBS. Dr. Feng Shao is the corresponding author. The research was supported by the 973 National High-Tech. Projects, the Beijing Municipal Government, China National Science Foundation, the Strategic Priority Research Program of the Chinese Academy of Sciences and Howard Hughes Medical Institute in the States, and carried out at National Institute of Biological Sciences, Beijing.
