Abstract:Accuracy in translation of the genetic code into proteins depends upon correct tRNA–mRNA recognition in the context of the ribosome. In human tRNAUUULys,3 three modified bases are present in the anticodon stem–loop—2-methylthio-N6-threonylcarbamoyladenosine at position 37 (ms2t6A37), 5-methoxycarbonylmethyl-2-thiouridine at position 34 (mcm5s2U34) and pseudouridine (ψ) at position 39—two of which, ms2t6A37 and mcm5s2U34, are required to achieve wild-type binding activity of wild-type human tRNAUUULys,3 [C. Yar… Show more
“…Previous molecular dynamics modeling and NMR spectrometry experiments had indicated that lengthy modifications at position-5 of U's could freeze the conformation of the nucleotide and dictate the hydrogen bonding geometry Durant et al 2005;McCrate et al 2006;Weixlbaumer et al 2007;Vendeix et al 2008). Results from NMR studies of mononucleoside 59-phosphates had indicated that the mnm 5 -modification formed a hydrogen bond to the nucleotide's 59-phosphate and that this additional bond stabilized the ribose C39-endo sugar pucker.…”
The maturation of RNAs includes site-specific post-transcriptional modifications that contribute significantly to hydrogen bond formation within RNA and between different RNAs, especially in formation of mismatch base pairs. Thus, an understanding of the geometry and strength of the base-pairing of modified ribonucleoside 59-monophosphates, previously not defined, is applicable to investigations of RNA structure and function and of the design of novel RNAs. The geometry and free energies of base-pairings were calculated in aqueous solution under neutral conditions with AMBER force fields and molecular dynamics simulations (MDSs). For example, unmodified uridines were observed to bind to uridine and cytidine with significant stability, but the ribose C19-C19 distances were far short (;8.9 Å ) of distances observed for canonical A-form RNA helices. In contrast, 5-oxyacetic acid uridine, known to bind adenosine, wobble to guanosine, and form mismatch base pairs with uridine and cytidine, bound adenosine and guanosine with geometries and energies comparable to an unmodified uridine. However, the 5-oxyacetic acid uridine base paired to uridine and cytidine with a C19-C19 distance comparable to that of an A-form helix, ;11 Å , when a H 2 O molecule migrated between and stably hydrogen bonded to both bases. Even in formation of canonical base pairs, intermediate structures with a second energy minimum consisted of transient H 2 O molecules forming hydrogen bonded bridges between the two bases. Thus, MDS is predictive of the effects of modifications, H 2 O molecule intervention in the formation of base-pair geometry, and energies that are important for native RNA structure and function.
“…Previous molecular dynamics modeling and NMR spectrometry experiments had indicated that lengthy modifications at position-5 of U's could freeze the conformation of the nucleotide and dictate the hydrogen bonding geometry Durant et al 2005;McCrate et al 2006;Weixlbaumer et al 2007;Vendeix et al 2008). Results from NMR studies of mononucleoside 59-phosphates had indicated that the mnm 5 -modification formed a hydrogen bond to the nucleotide's 59-phosphate and that this additional bond stabilized the ribose C39-endo sugar pucker.…”
The maturation of RNAs includes site-specific post-transcriptional modifications that contribute significantly to hydrogen bond formation within RNA and between different RNAs, especially in formation of mismatch base pairs. Thus, an understanding of the geometry and strength of the base-pairing of modified ribonucleoside 59-monophosphates, previously not defined, is applicable to investigations of RNA structure and function and of the design of novel RNAs. The geometry and free energies of base-pairings were calculated in aqueous solution under neutral conditions with AMBER force fields and molecular dynamics simulations (MDSs). For example, unmodified uridines were observed to bind to uridine and cytidine with significant stability, but the ribose C19-C19 distances were far short (;8.9 Å ) of distances observed for canonical A-form RNA helices. In contrast, 5-oxyacetic acid uridine, known to bind adenosine, wobble to guanosine, and form mismatch base pairs with uridine and cytidine, bound adenosine and guanosine with geometries and energies comparable to an unmodified uridine. However, the 5-oxyacetic acid uridine base paired to uridine and cytidine with a C19-C19 distance comparable to that of an A-form helix, ;11 Å , when a H 2 O molecule migrated between and stably hydrogen bonded to both bases. Even in formation of canonical base pairs, intermediate structures with a second energy minimum consisted of transient H 2 O molecules forming hydrogen bonded bridges between the two bases. Thus, MDS is predictive of the effects of modifications, H 2 O molecule intervention in the formation of base-pair geometry, and energies that are important for native RNA structure and function.
“…The final data are an average of the 10 scans (Figure 5B). The molecular dynamics simulation (MDS) were performed by following standard published protocol (23) with the exception of using a truncated octahedral TIP3P water box (24). Figure 5.Thermal denaturations and circular dichroism spectra of the -Ψ 27 and the -Ψ 27 ;f 5 C 34 .…”
Human mitochondrial methionine transfer RNA (hmtRNAMetCAU) has a unique post-transcriptional modification, 5-formylcytidine, at the wobble position-34 (f5C34). The role of this modification in (hmtRNAMetCAU) for the decoding of AUA, as well as AUG, in both the peptidyl- and aminoacyl-sites of the ribosome in either chain initiation or chain elongation is still unknown. We report the first synthesis and analyses of the tRNA's anticodon stem and loop domain containing the 5-formylcytidine modification. The modification contributes to the tRNA's anticodon domain structure, thermodynamic properties and its ability to bind codons AUA and AUG in translational initiation and elongation.
“…All systems were treated with periodic boundary conditions, and the equilibration protocol of Cheatham and Kollman 103 was performed similar to previous nucleic acid simulations. 85,104 Trajectories were propagated with a 2.0-fs time step with statistics collected every 1.0 ps. Equilibration times were established for the Rev-RRE, RSG 1.2-RRE IIB, and Rev-RRE complexes under different salt concentrations before statistics were collected over 10.0 ns production runs ( Table 1).…”
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