Changing nucleotide specificity of the DEAD-box helicase Hera abrogates communication between the Q-motif and the P-loop
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Julian Strohmeier
Abstract
DEAD-box proteins disrupt or remodel RNA and protein/RNA complexes at the expense of ATP. The catalytic core is composed of two flexibly connected RecA-like domains. The N-terminal domain contains most of the motifs involved in nucleotide binding and serves as a minimalistic model for helicase/nucleotide interactions. A single conserved glutamine in the so-called Q-motif has been suggested as a conformational sensor for the nucleotide state. To reprogram the Thermus thermophilus RNA helicase Hera for use of oxo-ATP instead of ATP and to investigate the sensor function of the Q-motif, we analyzed helicase activity of Hera Q28E. Crystal structures of the Hera N-terminal domain Q28E mutant (TthDEAD_Q28E) in apo- and ligand-bound forms show that Q28E does change specificity from adenine to 8-oxoadenine. However, significant structural changes accompany the Q28E mutation, which prevent the P-loop from adopting its catalytically active conformation and explain the lack of helicase activity of Hera_Q28E with either ATP or 8-oxo-ATP as energy sources. 8-Oxo-adenosine, 8-oxo-AMP, and 8-oxo-ADP weakly bind to TthDEAD_Q28E but in non-canonical modes. These results indicate that the Q-motif not only senses the nucleotide state of the helicase but could also stabilize a catalytically competent conformation of the P-loop and other helicase signature motifs.
©2011 by Walter de Gruyter Berlin New York
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Articles in the same Issue
- Guest Editorial
- Highlight: Mechanisms of RNA-mediated regulation
- HIGHLIGHT: 62ND MOSBACH COLLOQUIUM OF THE GBM ‘MECHANISMS OF RNA-MEDIATED REGULATION’
- Clustered regularly interspaced short palindromic repeats (CRISPRs): the hallmark of an ingenious antiviral defense mechanism in prokaryotes
- Regulatory RNAs in cyanobacteria: developmental decisions, stress responses and a plethora of chromosomally encoded cis-antisense RNAs
- RNA-dependent RNA polymerases in RNA silencing
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- Role of unique basic residues in cytotoxic, antibacterial and antiparasitic activities of human eosinophil cationic protein
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- Organelle stress-induced activating transcription factor-3 downregulates low-density lipoprotein receptor expression in Sk-Hep1 human liver cells
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