Formamide-Based Prebiotic Synthesis of Nucleobases: A Kinetically Accessible Reaction Route

Synthesis of nucleobases in nonaqueous environments is an alternative way for the emergence of terrestrial life, which could solve the fundamental problem connected to the hydrolytic instability of nucleic acid components in an aqueous environment. In this contribution, we present a plausible reaction route for the prebiotic synthesis of nucleobases in formamide, which does not require participation of the formamide trimer and aminoimidazole-carbonitrile intermediates. The computed activation energy of the proposed pathway is noticeably higher than that of the HCN-based synthetic route, but it is still feasible under the experimental conditions of the Saladino synthesis. We show that, albeit both the pyrimidine and purine ring formation utilizes the undissociated form of formamide, the dehydration product of formamide, HCN, may also play a key role in the mechanism. The rate determining step of the entire reaction path is the cyclization of the diaza-pentanimine precursor. The subsequent formation of the imidazole ring proceeds with a moderate activation energy. Our calculations thus demonstrate that the experimentally suggested reaction path without the involvement of aminoimidazole-carbonitrile intermediates is also a viable alternative for the nonaqueous synthesis of nucleobases

Comment on “Computational Model for Predicting Experimental RNA and DNA Nearest-Neighbor Free Energy Rankings”

Comment on “Computational Model for Predicting Experimental RNA and DNA Nearest-Neighbor Free Energy Rankings

Derivation of Reliable Geometries in QM Calculations of DNA Structures: Explicit Solvent QM/MM and Restrained Implicit Solvent QM Optimizations of G‑Quadruplexes

Modern dispersion-corrected DFT methods have made it possible to perform reliable QM studies on complete nucleic acid (NA) building blocks having hundreds of atoms. Such calculations, although still limited to investigations of potential energy surfaces, enhance the portfolio of computational methods applicable to NAs and offer considerably more accurate intrinsic descriptions of NAs than standard MM. However, in practice such calculations are hampered by the use of implicit solvent environments and truncation of the systems. Conventional QM optimizations are spoiled by spurious intramolecular interactions and severe structural deformations. Here we compare two approaches designed to suppress such artifacts: partially restrained continuum solvent QM and explicit solvent QM/MM optimizations. We report geometry relaxations of a set of diverse double-quartet guanine quadruplex (GQ) DNA stems. Both methods provide neat structures without major artifacts. However, each one also has distinct weaknesses. In restrained optimizations, all errors in the target geometries (i.e., low-resolution X-ray and NMR structures) are transferred to the optimized geometries. In QM/MM, the initial solvent configuration causes some heterogeneity in the geometries. Nevertheless, both approaches represent a decisive step forward compared to conventional optimizations. We refine earlier computations that revealed sizable differences in the relative energies of GQ stems computed with AMBER MM and QM. We also explore the dependence of the QM/MM results on the applied computational protocol

Electron-Driven Proton Transfer Along H₂O Wires Enables Photorelaxation of πσ* States in Chromophore–Water Clusters

The fates of photochemically formed πσ* states are one of the central issues in photobiology due to their significant contribution to the photostability of biological matter, formation of hydrated electrons, and the phenomenon of photoacidity. Nevertheless, our understanding of the underlying molecular mechanisms in aqueous solution is still incomplete. In this paper, we report on the results of nonadiabatic photodynamics simulations of microhydrated 2-aminooxazole molecule employing algebraic diagrammatic construction to the second order. Our results indicate that electron-driven proton transfer along H₂O wires induces the formation of πσ*/S₀ state crossing and provides an effective deactivation channel. Because we recently have identified a similar channel for 4-aminoimidazole-5-carbonitrile [Szabla, R.; Phys. Chem. Chem. Phys. 2014, 16, 17617−17626], we conclude this mechanism may be quite common to all heterocyclic compounds with low-lying πσ* states

Insight into G-DNA Structural Polymorphism and Folding from Sequence and Loop Connectivity through Free Energy Analysis

The lengths of G-tracts and their connecting loop sequences determine G-quadruplex folding and stability. Complete understanding of the sequence–structure relationships remains elusive. Here, single-loop G-quadruplexes were investigated using explicit solvent molecular dynamics (MD) simulations to characterize the effect of loop length, loop sequence, and G-tract length on the folding topologies and stability of G-quadruplexes. Eight loop types, including different variants of lateral, diagonal, and propeller loops, and six different loop sequences [d0 (i.e., no intervening residues in the loop), dT, dT₂, dT₃, dTTA, and dT₄] were considered through MD simulation and free energy analysis. In most cases the free energetic estimates agree well with the experimental observations. The work also provides new insight into G-quadruplex folding and stability. This includes reporting the observed instability of the left propeller loop, which extends the rules for G-quadruplex folding. We also suggest a plausible explanation why human telomere sequences predominantly form hybrid-I and hybrid-II type structures in K⁺ solution. Overall, our calculation results indicate that short loops generally are less stable than longer loops, and we hypothesize that the extreme stability of sequences with very short loops could possibly derive from the formation of parallel multimers. The results suggest that free energy differences, estimated from MD and free energy analysis with current force fields and simulation protocols, are able to complement experiment and to help dissect and explain loop sequence, loop length, and G-tract length and orientation influences on G-quadruplex structure

Comparative Assessment of Different RNA Tetranucleotides from the DFT-D3 and Force Field Perspective

Classical force field (FF) molecular dynamics (MD) simulations of RNA tetranucleotides have substantial problems in reproducing conformer populations indicated by NMR experiments. To provide more information about the possible sources of errors, we performed quantum mechanical (QM, TPSS-D3/def2-TZVP) and molecular mechanics (MM, AMBER parm99bsc0+χ_OL3) calculations of different r­(CCCC), r­(GACC), and r­(UUUU) conformers obtained from explicit solvent MD simulations. Solvent effects in the static QM and MM calculations were mimicked using implicit solvent models (COSMO and Poisson–Boltzmann, respectively). The comparison of QM and MM geometries and energies revealed that the two methodologies provide qualitatively consistent results in most of the cases. Even though we found some differences, these were insufficient to indicate any systematic corrections of the RNA FF terms that could improve the performance of classical MD in simulating tetranucleotides. On the basis of these findings, we inferred that the overpopulation of intercalated conformers in the MD simulations of RNA tetramers, which were not observed experimentally, might be predominantly caused by imbalanced water–solvent and water–water interactions. Apart from the large-scale QM calculations performed to assess the performance of the AMBER FF, a representative spectrum of faster QM methods was tested

Are Waters around RNA More than Just a Solvent? – An Insight from Molecular Dynamics Simulations

Hydrating water molecules are believed to be an inherent part of the RNA structure and have a considerable impact on RNA conformation. However, the magnitude and mechanism of the interplay between water molecules and the RNA structure are still poorly understood. In principle, such hydration effects can be studied by molecular dynamics (MD) simulations. In our recent MD studies, we observed that the choice of water model has a visible impact on the predicted structure and structural dynamics of RNA and, in particular, has a larger effect than type, parametrization, and concentration of the ions. Furthermore, the water model effect is sequence dependent and modulates the sequence dependence of A-RNA helical parameters. Clearly, the sensitivity of A-RNA structural dynamics to the water model parametrization is a rather spurious effect that complicates MD studies of RNA molecules. These results nevertheless suggest that the sequence dependence of the A-RNA structure, usually attributed to base stacking, might be driven by the structural dynamics of specific hydration. Here, we present a systematic MD study that aimed to (i) clarify the atomistic mechanism of the water model sensitivity and (ii) discover whether and to what extent specific hydration modulates the A-RNA structural variability. We carried out an extended set of MD simulations of canonical A-RNA duplexes with TIP3P, TIP4P/2005, TIP5P, and SPC/E explicit water models and found that different water models provided a different extent of water bridging between 2′-OH groups across the minor groove, which in turn influences their distance and consequently also inclination, roll, and slide parameters. Minor groove hydration is also responsible for the sequence dependence of these helical parameters. Our simulations suggest that TIP5P is not optimal for RNA simulations

MD and QM/MM Study of the Quaternary HutP Homohexamer Complex with mRNA, l‑Histidine Ligand, and Mg²⁺

The HutP protein from <i>B. subtilis</i> regulates histidine metabolism by interacting with an antiterminator mRNA hairpin in response to the binding of l-histidine and Mg²⁺. We studied the functional ligand-bound HutP hexamer complexed with two mRNAs using all-atom microsecond-scale explicit-solvent MD simulations performed with the Amber force fields. The experimentally observed protein-RNA interface exhibited good structural stability in the simulations with the exception of some fluctuations in an unusual adenine-threonine interaction involving two closely spaced H-bonds. We further investigated this interaction by comparing QM/MM and MM optimizations, using the QM region comprising almost 350 atoms described at the DFT-D3 level. The QM/MM method clearly improved the adenine-threonine interaction compared to MM, especially when the X–H bond lengths were frozen during the MM optimization to mimic the use of SHAKE in the MD simulations. Thus, both the MM approximation and the use of SHAKE can compromise the description of H-bonds at protein-RNA interfaces. The simulations also revealed a notable Mg²⁺-parameter dependence in the behavior of the ligand-binding pocket (LBP). With the SPC/E water model, the 12–6 Åqvist and Li&Merz parameters provided an entirely stable LBP structure, but the 12–6 Allnér and 12–6–4 Li&Merz parametrizations resulted in a progressive loss of direct nitrogen–Mg²⁺ LBP coordination. The Allnér and Li&Merz 12–6 parametrizations were also tested with the TIP3P water model; the LBP was destabilized in both cases. This illustrates the difficulty of consistently describing different Mg²⁺ interactions using nonpolarizable force fields. Overall, the simulations support the hypothesis that HutP protein becomes fully structured upon ligand binding. Subsequent RNA binding does not affect the protein structure, in keeping with the mechanism inferred from experimental structures

MD and QM/MM Study of the Quaternary HutP Homohexamer Complex with mRNA, l‑Histidine Ligand, and Mg²⁺

The HutP protein from <i>B. subtilis</i> regulates histidine metabolism by interacting with an antiterminator mRNA hairpin in response to the binding of l-histidine and Mg²⁺. We studied the functional ligand-bound HutP hexamer complexed with two mRNAs using all-atom microsecond-scale explicit-solvent MD simulations performed with the Amber force fields. The experimentally observed protein-RNA interface exhibited good structural stability in the simulations with the exception of some fluctuations in an unusual adenine-threonine interaction involving two closely spaced H-bonds. We further investigated this interaction by comparing QM/MM and MM optimizations, using the QM region comprising almost 350 atoms described at the DFT-D3 level. The QM/MM method clearly improved the adenine-threonine interaction compared to MM, especially when the X–H bond lengths were frozen during the MM optimization to mimic the use of SHAKE in the MD simulations. Thus, both the MM approximation and the use of SHAKE can compromise the description of H-bonds at protein-RNA interfaces. The simulations also revealed a notable Mg²⁺-parameter dependence in the behavior of the ligand-binding pocket (LBP). With the SPC/E water model, the 12–6 Åqvist and Li&Merz parameters provided an entirely stable LBP structure, but the 12–6 Allnér and 12–6–4 Li&Merz parametrizations resulted in a progressive loss of direct nitrogen–Mg²⁺ LBP coordination. The Allnér and Li&Merz 12–6 parametrizations were also tested with the TIP3P water model; the LBP was destabilized in both cases. This illustrates the difficulty of consistently describing different Mg²⁺ interactions using nonpolarizable force fields. Overall, the simulations support the hypothesis that HutP protein becomes fully structured upon ligand binding. Subsequent RNA binding does not affect the protein structure, in keeping with the mechanism inferred from experimental structures

Bioinformatics and Molecular Dynamics Simulation Study of L1 Stalk Non-Canonical rRNA Elements: Kink-Turns, Loops, and Tetraloops

The L1 stalk is a prominent mobile element of the large ribosomal subunit. We explore the structure and dynamics of its non-canonical rRNA elements, which include two kink-turns, an internal loop, and a tetraloop. We use bioinformatics to identify the L1 stalk RNA conservation patterns and carry out over 11.5 μs of MD simulations for a set of systems ranging from isolated RNA building blocks up to complexes of L1 stalk rRNA with the L1 protein and tRNA fragment. We show that the L1 stalk tetraloop has an unusual GNNA or UNNG conservation pattern deviating from major GNRA and YNMG RNA tetraloop families. We suggest that this deviation is related to a highly conserved tertiary contact within the L1 stalk. The available X-ray structures contain only UCCG tetraloops which in addition differ in orientation (<i>anti</i> vs <i>syn</i>) of the guanine. Our analysis suggests that the <i>anti</i> orientation might be a mis-refinement, although even the <i>anti</i> interaction would be compatible with the sequence pattern and observed tertiary interaction. Alternatively, the <i>anti</i> conformation may be a real substate whose population could be pH-dependent, since the guanine <i>syn</i> orientation requires protonation of cytosine in the tertiary contact. In absence of structural data, we use molecular modeling to explore the GCCA tetraloop that is dominant in bacteria and suggest that the GCCA tetraloop is structurally similar to the YNMG tetraloop. Kink-turn Kt-77 is unusual due to its 11-nucleotide bulge. The simulations indicate that the long bulge is a stalk-specific eight-nucleotide insertion into consensual kink-turn only subtly modifying its structural dynamics. We discuss a possible evolutionary role of helix H78 and a mechanism of L1 stalk interaction with tRNA. We also assess the simulation methodology. The simulations provide a good description of the studied systems with the latest bsc0χ_OL3 force field showing improved performance. Still, even bsc0χ_OL3 is unable to fully stabilize an essential sugar-edge H-bond between the bulge and non-canonical stem of the kink-turn. Inclusion of Mg²⁺ ions may deteriorate the simulations. On the other hand, monovalent ions can in simulations readily occupy experimental Mg²⁺ binding sites