Ribitol in Solution Is an Equilibrium of Asymmetric Conformations

Ribitol (C5H12O5), an acyclic sugar alcohol, is present on mammalian α-dystroglycan as a component of O-mannose glycan. In this study, we examine the conformation and dynamics of ribitol by database analysis, experiments, and computational methods. Database analysis reveals that the anti-conformation (180°) is populated at the C3–C4 dihedral angle, while the gauche conformation (±60°) is seen at the C2–C3 dihedral angle. Such conformational asymmetry was born out in a solid-state 13C-NMR spectrum of crystalline ribitol, where C1 and C5 signals are unequal. On the other hand, solution 13C-NMR has identical chemical shifts for C1 and C5. NMR 3J coupling constants and OH exchange rates suggest that ribitol is an equilibrium of conformations, under the influence of hydrogen bonds and/or steric hinderance. Molecular dynamics (MD) simulations allowed us to discuss such a chemically symmetric molecule, pinpointing the presence of asymmetric conformations evidenced by the presence of correlations between C2–C3 and C3–C4 dihedral angles. These findings provide a basis for understanding the dynamic structure of ribitol and the function of ribitol-binding enzymes.

Insights into the mismatch discrimination mechanism of Y-family DNA polymerase Dpo4

Nucleobases within DNA are attacked by reactive oxygen species to produce 7,8-dihydro-8-oxoguanine (oxoG) and 7,8-dihydro-8-oxoadenine (oxoA) as major oxidative lesions. The high mutagenicity of oxoG is attributed to the lesion’s ability to adopt syn-oxoG:anti-dA with Watson-Crick-like geometry. Recent studies have revealed that Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) inserts nucleotide opposite oxoA in an error-prone manner and accommodates syn-oxoA:anti-dGTP with Watson-Crick-like geometry, highlighting a promutagenic nature of oxoA. To gain further insights into the bypass of oxoA by Dpo4, we have conducted kinetic and structural studies of Dpo4 extending oxoA:dT and oxoA:dG by incorporating dATP opposite templating dT. The extension past oxoA:dG was ~5-fold less efficient than that past oxoA:dT. Structural studies revealed that Dpo4 accommodated dT:dATP base pair past anti-oxoA:dT with little structural distortion. In the Dpo4-oxoA:dG extension structure, oxoA was in an anti conformation and did not form hydrogen bonds with the primer terminus base. Unexpectedely, the dG opposite oxoA exited the primer terminus site and resided in an extrahelical site, where it engaged in minor groove contacts to the two immediate upstream bases. The extrahelical dG conformation appears to be induced by the stabilization of anti-oxoA conformation via bifurcated hydrogen bonds with Arg332. This unprecedented structure suggests that Dpo4 may use Arg332 to sense 8-oxopurines at the primer terminus site and slow the extension from the mismatch by promoting anti conformation of 8-oxopurines.

Promutagenic bypass of 7,8-dihydro-8-oxoadenine by translesion synthesis DNA polymerase Dpo4

Reactive oxygen species induced by ionizing radiation and metabolic pathways generate 7,8-dihydro-8-oxoguanine (oxoG) and 7,8-dihydro-8-oxoadenine (oxoA) as two major forms of oxidative damage. The mutagenicity of oxoG, which promotes G to T transversions, is attributed to the lesion’s conformational flexibility that enables Hoogsteen base pairing with dATP in the confines of DNA polymerases. The mutagenesis mechanism of oxoA, which preferentially causes A to C transversions, remains poorly characterized. While structures for oxoA bypass by human DNA polymerases are available, that of prokaryotic DNA polymerases have not been reported. Herein, we report kinetic and structural characterizations of Sulfolobus solfataricus Dpo4 incorporating a nucleotide opposite oxoA. Our kinetic studies show oxoA at the templating position reduces the replication fidelity by ~560-fold. The catalytic efficiency of the oxoA:dGTP insertion is ~300-fold greater than that of the dA:dGTP insertion, highlighting the promutagenic nature of oxoA. The relative efficiency of the oxoA:dGTP misincorporation is ~5-fold greater than that of the oxoG:dATP misincorporation, suggesting the mutagenicity of oxoA is comparable to that of oxoG. In the Dpo4 replicating base pair site, oxoA in the anti-conformation forms a Watson-Crick base pair with an incoming dTTP, while oxoA in the syn-conformation assumes Hoogsteen base pairing with an incoming dGTP, displaying the dual coding potential of the lesion. Within the Dpo4 active site, the oxoA:dGTP base pair adopts a Watson-Crick-like geometry, indicating Dpo4 influences the oxoA:dGTP base pair conformation. Overall, the results reported here provide insights into the miscoding properties of the major oxidative adenine lesion during translesion synthesis.

The hyperthermophilic partners Nanoarchaeum and Ignicoccus stabilize their tRNA T-loops via different but structurally equivalent modifications

The universal L-shaped tertiary structure of tRNAs is maintained with the help of nucleotide modifications within the D- and T-loops, and these modifications are most extensive within hyperthermophilic species. The obligate-commensal Nanoarchaeum equitans and its phylogenetically-distinct host Ignicoccus hospitalis grow physically coupled under identical hyperthermic conditions. We report here two fundamentally different routes by which these archaea modify the key conserved nucleotide U54 within their tRNA T-loops. In N. equitans, this nucleotide is methylated by the S-adenosylmethionine-dependent enzyme NEQ053 to form m5U54, and a recombinant version of this enzyme maintains specificity for U54 in Escherichia coli. In N. equitans, m5U54 is subsequently thiolated to form m5s2U54. In contrast, I. hospitalis isomerizes U54 to pseudouridine prior to methylating its N1-position and thiolating the O4-position of the nucleobase to form the previously uncharacterized nucleotide m1s4Ψ. The methyl and thiol groups in m1s4Ψ and m5s2U are presented within the T-loop in a spatially identical manner that stabilizes the 3′-endo-anti conformation of nucleotide-54, facilitating stacking onto adjacent nucleotides and reverse-Hoogsteen pairing with nucleotide m1A58. Thus, two distinct structurally-equivalent solutions have evolved independently and convergently to maintain the tertiary fold of tRNAs under extreme hyperthermic conditions.

Meso-1,4-phenylene bridged nickel norcorrole dimer

Nickel norcorrole dimer 3 linked with a phenylene bridge was prepared by a nickel-mediated homolytic cross-coupling of meso-phenylene bisdipyrrin nickel(II) dimer 2b. The structures of 2b and 3 were elucidated by X-ray diffraction analysis. Each nickel norcorrole fraction of the nickel norcorrole dimer 3 exposed a bowl-like structure, which conserved anti-conformation. Structural distinctions between the monomeric and dimeric nickel norcorroles were established by vibrational spectroscopic analysis and assessed with density functional theory calculations. Enhanced flexibility of the norcorrole planes in solution states of monomeric nickel norcorrole was compromised to that of bowl-shaped norcorroles in the solid states of nickel norcorrole dimer 3, where C[Formula: see text] was the formulated conformation. The deformation of the planarity derived from the objection of laser pulses effectuated vibration shifts of specific Raman-active motions toward a higher-frequency region, as associating with a paratropic nature of the [Formula: see text]-electron delocalization circuit of norcorrole. Computational simulation exposed a reliable drift of the Raman frequencies.

In Silico Acetylene [2+2+2] Cycloadditions Catalyzed by Rh/Cr Indenyl Fragments

Metal-catalyzed alkyne [2+2+2] cycloadditions provide a variety of substantial aromatic compounds of interest in the chemical and pharmaceutical industries. Herein, the mechanistic aspects of the acetylene [2+2+2] cycloaddition mediated by bimetallic half-sandwich catalysts [Cr(CO)3IndRh] (Ind = (C9H7)−, indenyl anion) are investigated. A detailed exploration of the potential energy surfaces (PESs) was carried out to identify the intermediates and transition states, using a relativistic density functional theory (DFT) approach. For comparison, monometallic parent systems, i.e., CpRh (Cp = (C5H5)−, cyclopentadienyl anion) and IndRh, were included in the analysis. The active center is the rhodium nucleus, where the [2+2+2] cycloaddition occurs. The coordination of the Cr(CO)3 group, which may be in syn or anti conformation, affects the energetics of the catalytic cycle as well as the mechanism. The reaction and activation energies and the turnover frequency (TOF) of the catalytic cycles are rationalized, and, in agreement with the experimental findings, our computational analysis reveals that the presence of the second metal favors the catalysis.

Synthesis and Light-Induced Actuation of Photo-Labile 2-Pyridyl-1,2,3-Triazole Ru(bis-bipyridyl) Appended Ferrocene Rotors

To realise useful control over molecular motion in the future an extensive toolbox of both actionable molecules and stimuli-responsive units must be developed. Previously, our laboratory has reported 1,1′-disubstituted ferrocene (Fc) rotor units which assume a contracted/π-stacked conformation until complexation of cationic metal ions causes rotation about the Ferrocene (Fc) molecular ‘ball-bearing’. Herein, we explore the potential of using the photochemical ejection of [Ru(2,2′-bipyridyl)2]2+ units as a stimulus for the rotational contraction of new ferrocene rotor units. Fc rotors with both ‘regular’ and ‘inverse’ 2-pyridyl-1,2,3-triazole binding pockets and their corresponding [Ru(2,2′-bipyridyl)2]2+ complexes were synthesised. The rotors and complexes were characterised using nuclear magnetic resonance (NMR) and ultraviolet (UV)-visible spectroscopies, Electro-Spray Ionisation Mass Spectrometry (ESI–MS), and electrochemistry. The 1,1′-disubstituted Fc ligands were shown to π-stack both in solution and solid state. Density Functional Theory (DFT) calculations (CAM-B3LYP/6-31G(d)) support the notion that complexation to [Ru(2,2′-bipyridyl)2]2+ caused a rotation from the syn- to the anti-conformation. Upon photo-irradiation with UV light (254 nm), photo-ejection of the [Ru(2,2′-bipyridyl)2(CH3CN)2]2+ units in acetonitrile was observed. The re-complexation of the [Ru(2,2′-bipyridyl)2]2+ units could be achieved using acetone as the reaction solvent. However, the process was exceedingly slowly. Additionally, the Fc ligands slowly decomposed when exposed to UV irradiation meaning that only one extension and contraction cycle could be completed.