Thermodynamics of the pseudo‐knot in helix 18 of 16S ribosomal RNA

Abstract

A fragment of E. coli 16S rRNA formed by nucleotides 500 to 545 is termed helix 18. Nucleotides 505-507 and 524-526 form a pseudo-knot and its distortion affects ribosome function. Helix 18 isolated from the ribosome context is thus an interesting fragment to investigate the structural properties and folding of RNA with pseudo-knots. With all-atom molecular dynamics simulations, spectroscopic and gel electrophoresis experiments, we investigated thermodynamics of helix 18, with a focus on its pseudo-knot. In solution studies at ambient conditions we observed dimerization of helix 18. We proposed that the loop, containing nucleotides forming the pseudo-knot, interacts with another monomer of helix 18. The native dimer is difficult to break but introducing mutations in the pseudo-knot indeed assured a monomeric form of helix 18. Molecular dynamics simulations at 310 K confirmed the stability of the pseudo-knot but at elevated temperatures this pseudo-knot was the first part of helix 18 to lose the hydrogen bond pattern. To further determine helix 18 stability, we analyzed the interactions of helix 18 with short oligomers complementary to a nucleotide stretch containing the pseudo-knot. The formation of higher-order structures by helix 18 impacts hybridization efficiency of peptide nucleic acid and 2'-O methyl RNA oligomers.

Quantitative comparison between sub-millisecond time resolution single-molecule FRET measurements and 10-second molecular simulations of a biosensor protein

“…When data from these techniques are directly compared to MD simulations, the time scale of the simulations is often not considered critical, so long as sampling is sufficient to avoid equilibration artifacts . Beyond these techniques, MD simulations can provide atomistic insights into structural dynamics for a multitude of experimental approaches [39][40][41][42][43][44][45][46]. Yet, close quantitative comparisons between in vitro experimental data and in silico simulations at commensurate time scales are relatively rare.…”

Modulating protein translation through proposed mRNA-rRNA base pairing

“…This suggests that the conformation of loop in reconstituted ribosome structures may have a relatively high free energy of folding. This idea is supported by molecular dynamics simulations of this region that show that the pseudoknot formed by nucleotides G524-C526 was the first region of helix 18 of the 16S rRNA to become disordered in melting experiments (Wojciechowska, Dudek, & Trylska, 2018). G529, C528, and G527 are uniquely positioned closest to the mRNA entrance tunnel and would therefore have the greatest potential to base pair to mRNA sequences.…”