Abstract
Argonaute proteins in combination with short microRNA (miRNAs) can target mRNA molecules for translation inhibition or degradation and play a key role in many regulatory processes. The miRNAs act as guide RNAs that associate with Argonaute and the complementary mRNA target region. The complex formation results in activation of Argonaute and specific cleavage of the target mRNA. Both the binding and activation processes involve essential domain rearrangements of functional importance. For the Thermus Thermophilus Argonaute (TtAgo) system guide-bound (binary) and guide/target-bound (ternary) complexes are known but how the binding of guide and target mediate domain movements is still not understood. We have studied the Argonaute domain motion in apo and guide/target bound states using Molecular Dynamics simulations and a Hamiltonian replica exchange (H-REMD) method that employs a specific biasing potential to accelerate domain motions. The H-REMD technique indicates sampling of a much broader distribution of domain arrangements both in the apo as well as binary and ternary complexes compared to regular MD simulations. In the apo state domain arrangements corresponding to more compact (closed) states are mainly sampled which undergo an opening upon guide and guide/target binding. Whereas only limited overlap in domain geometry between apo and bound states was found, a larger similarity in the domain distribution is observed for the simulations of binary and ternary complexes. Comparative simulations on ternary complexes with 15 or 16 base pairs (bp) formed between guide and target strands (instead of 14) resulted in dissociation of the 3'-guide strand from the PAZ domain and domain rearrangement. This agrees with the experimental observation that guide-target pairing beyond 14 bps is required for activation and gives a mechanistic explanation for the experimentally observed activation process.
Highlights
The Argonautes constitute a family of proteins that are involved in both transcriptional and post-transcriptional gene regulatory mechanisms [1–3]
Argonaute domain motion studied with Hamiltonian replica-exchange molecular dynamics (H-REMD) simulations
Argonautes are key components of the RNA-induced Silencing Complex (RISC), which drives silencing of a target messenger RNA via degradation or repression
Summary
The Argonautes constitute a family of proteins that are involved in both transcriptional and post-transcriptional gene regulatory mechanisms [1–3]. They are present in all forms of life, sharing a well conserved tertiary structure, despite their limited sequence similarity [4–6]. The regulation of gene expression via miRNAs is different from that of the siRNA, as the latter binds to a fully complementary target mRNA and prompts specific gene silencing, while the former can regulate multiple mRNAs. Prokaryotic Argonautes, on the other hand, participate in gene regulation by binding to single-stranded DNAs that guide them towards target DNAs or RNAs that are either cleaved or repressed [7–10]. Bacterial Argonaute protects its host against invasive genomic elements through directly targeting foreign DNA molecules [9,11]
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