Influence of 2'-Modifications (O-Methylation, Fluorination, and Stereochemical Inversion) on the Base Pairing Energies of Protonated Cytidine Nucleoside Analogue Base Pairs: Implications for the Stabilities of i-Motif Structures.

Naturally occurring and chemically engineered modifications are among the most powerful strategies explored for fine-tuning the conformational characteristics and intrinsic stability of nucleic acids topologies. Modifications at the 2'-position of the ribose or 2'-deoxyribose moieties differentiate nucleic acid structures and have a significant impact on their electronic properties and base-pairing interactions. 2'-O-Methylation, a common post-transcriptional modification of tRNA, is directly involved in modulating specific anticodon-codon base-pairing interactions. 2'-Fluorinated and arabino nucleosides possess novel and beneficial medicinal properties and find use as therapeutics for treating viral diseases and cancer. However, the potential to deploy 2'-modified cytidine chemistries for tuning i-motif stability is largely unknown. To address this knowledge gap, the effects of 2'-modifications including O-methylation, fluorination, and stereochemical inversion on the base-pairing interactions of protonated cytidine nucleoside analogue base pairs, the core stabilizing interactions of i-motif structures, are examined using complementary threshold collision-induced dissociation techniques and computational methods. The 2'-modified cytidine nucleoside analogues investigated here include 2'-O-methylcytidine, 2'-fluoro-2'-deoxycytidine, arabinofuranosylcytosine, 2'-fluoro-arabinofuranosylcytosine, and 2',2'-difluoro-2'-deoxycytidine. All five 2'-modifications examined here are found to enhance the base-pairing interactions relative to the canonical DNA and RNA cytidine nucleosides with the greatest enhancements arising from 2'-O-methylation and 2',2'-difluorination, suggesting that these modifications should well be tolerated in the narrow grooves of i-motif conformations.

The Ideal Physical Therapist from the Perspective of Individuals With Limb Loss.

BACKGROUND: Effective rehabilitation after limb loss is necessary to maximize function and promote independence. Physical therapists (PT) are one of the primary drivers of the rehabilitation process. While general physical therapy knowledge and abilities have been shown to be important to the rehabilitation process, it is unclear what individuals with limb loss value in their PT's. OBJECTIVE: The purpose of this study was to understand the elements that define an ideal PT from the perspective of individuals with limb loss. METHODOLOGY: Mixed-method design consisting of a 20-item web-based survey and semi-structured interviews that were administered to individuals 18 years or older, who spoke English, and had a history of lower limb loss. FINDINGS: Individuals with limb loss describe an ideal PT as promoting a therapeutic alliance, having specialized knowledge, and collaborating with a prosthetist. Knowledge of the PT as it relates to limb loss was found to be both the greatest facilitator and barrier to the rehabilitation process. CONCLUSION: From the perspective of those with limb loss, an ideal PT promotes a strong therapeutic alliance through communication, has specialized knowledge when it comes to the limb loss rehabilitation process, and collaborates with the prosthetist to problem-solve throughout the rehabilitation process.

Structural Determination of Lysine-Linked Cisplatin Complexes via IRMPD Action Spectroscopy: NNs and NO- Binding Modes of Lysine to Platinum Coexist.

Despite its success as an anticancer drug, cisplatin suffers from resistance and produces side effects. To overcome these limitations, amino-acid-linked cisplatin analogues have been investigated. Lysine-linked cisplatin, Lysplatin, (Lys)PtCl2, exhibited outstanding reactivity toward DNA and RNA that differs from that of cisplatin. To gain insight into its differing reactivity, the structure of Lysplatin is examined here using infrared multiple photon dissociation (IRMPD) action spectroscopy. To probe the influence of the local chemical environment on structure, the deprotonated and sodium-cationized Lysplatin complexes are examined. Electronic structure calculations are performed to explore possible modes of binding of Lys to Pt, their relative stabilities, and to predict their infrared spectra. Comparisons of the measured IRMPD and predicted IR spectra elucidate the structures contributing to the experimental spectra. Coexistence of two modes of binding of Lys to Pt is found where Lys binds via the backbone and side-chain amino nitrogen atoms, NNs, or to the backbone amino and carboxylate oxygen atoms, NO-. Glycine-linked cisplatin and arginine-linked cisplatin complexes have previously been found to bind only via the NO- binding mode. Present results suggest that the NNs binding conformers may be key to the outstanding reactivity of Lysplatin toward DNA and RNA.

5-Halogenation of Uridine Suppresses Protonation-Induced Tautomerization and Enhances Glycosidic Bond Stability of Protonated Uridine: Investigations via IRMPD Action Spectroscopy, ER-CID Experiments, and Theoretical Calculations.

Uridine (Urd), a canonical nucleoside of RNA, is the most commonly modified nucleoside among those that occur naturally. Uridine has also been an important target for the development of modified nucleoside analogues for pharmaceutical applications. In this work, the effects of 5-halogenation of uracil on the structures and glycosidic bond stabilities of protonated uridine nucleoside analogues are examined using tandem mass spectrometry and computational methods. Infrared multiple photon dissociation (IRMPD) action spectroscopy experiments and theoretical calculations are performed to probe the structural influences of these modifications. Energy-resolved collision-induced dissociation experiments along with survival yield analyses are performed to probe glycosidic bond stability. The measured IRMPD spectra are compared to linear IR spectra predicted for the stable low-energy conformations of these species computed at the B3LYP/6-311+G(d,p) level of theory to determine the conformations experimentally populated. Spectral signatures in the IR fingerprint and hydrogen-stretching regions allow the 2,4-dihydroxy protonated tautomers (T) and O4- and O2-protonated conformers to be readily differentiated. Comparisons between the measured and predicted spectra indicate that parallel to findings for uridine, both T and O4-protonated conformers of the 5-halouridine nucleoside analogues are populated, whereas O2-protonated conformers are not. Variations in yields of the spectral signatures characteristic of the T and O4-protonated conformers indicate that the extent of protonation-induced tautomerization is suppressed as the size of the halogen substituent increases. Trends in the energy-dependence of the survival yield curves find that 5-halogenation strengthens the glycosidic bond and that the enhancement in stability increases with the size of the halogen substituent.

Influence of 5-Halogenation on the Base-Pairing Energies of Protonated Cytidine Nucleoside Analogue Base Pairs: Implications for the Stabilities of Synthetic i-Motif Structures for DNA Nanotechnology Applications.

DNA nanotechnology has been employed to develop devices based on i-motif structures. The protonated cytosine-cytosine base pairs that stabilize i-motif conformations are favored under slightly acidic conditions. This unique property has enabled development of the first DNA molecular motor driven by pH changes. The ability to alter the stability and pH transition range of such DNA molecular motors is desirable. Understanding how i-motif structures are influenced by modifications, and which modifications enhance stability and/or affect the pH characteristics, are therefore of great interest. Here, the influence of 5-halogenation of the cytosine nucleobases on the base pairing of protonated cytidine nucleoside analogue base pairs is examined using complementary threshold collision-induced dissociation techniques and computational methods. The nucleoside analogues examined here include the 5-halogenated forms of the canonical DNA and RNA cytidine nucleosides. Comparisons among these systems and to the analogous canonical base pairs previously examined enable the influence of 5-halogenation and the 2'-hydroxy substituent on the base pairing to be elucidated. 5-Halogenation of the cytosine nucleobases is found to enhance the strength of base pairing of DNA base pairs and generally weakens the base pairing for RNA base pairs. Trends in the strength of base pairing indicate that both inductive and polarizability effects influence the strength of base pairing. Overall, the present results suggest that 5-halogenation, and in particular, 5-fluorination and 5-iodination, provide effective means of stabilizing DNA i-motif conformations for applications in nanotechnology, whereas only 5-iodination is effective for stabilizing RNA i-motif conformations but the enhancement in stability is less significant.

Off-Pump Coronary Artery Bypass Grafting; is it Still Relevant?

Off-pump Coronary Artery Bypass Grafting (OPCAB) experienced a resurgence in the 1980s -2000s and developed steadily with improvement of the instrumentation and techniques. However questions about graft patency and long-term survival of OPCAB patients still exist. This review attempts to explore the current relevance of OPCAB.

Infrared multiple photon dissociation action spectroscopy of protonated unsymmetrical dimethylhydrazine and proton-bound dimers of hydrazine and unsymmetrical dimethylhydrazine.

The gas-phase structures of protonated unsymmetrical 1,1-dimethylhydrazine (UDMH) and the proton-bound dimers of UDMH and hydrazine are examined by infrared multiple photon dissociation (IRMPD) action spectroscopy utilizing light generated by a free electron laser and an optical parametric oscillator laser system. To identify the structures present in the experimental studies, the measured IRMPD spectra are compared to spectra calculated at the B3LYP-GD3BJ/6-311+G(d,p) level of theory. These comparisons show that protonated UDMH binds the proton at the methylated nitrogen atom (α) with two low-lying α conformers probably being populated. For (UDMH)2H+, the proton is shared between the methylated nitrogen atoms with several low-lying α conformers likely to be populated. Higher-lying conformers of (UDMH)2H+ in which the proton is shared between α and ß (unmethylated) nitrogen atoms cannot be ruled out on the basis of the IRPMD spectrum. For (N2H4)2H+, there are four low-lying conformers that all reproduce the IRMPD spectrum reasonably well. As hydrazine and UDMH see usage as fuels for rocket engines, such spectra are potentially useful as a means of remotely monitoring rocket launches, especially in cases of unsuccessful launches where environmental hazards need to be assessed.

1-Alkyl-3-methylimidazolium cation binding preferences in hexafluorophosphate ionic liquid clusters determined using competitive TCID measurements and theoretical calculations.

Ionic liquids (ILs) exhibit unique properties that have led to their development and widespread use for a variety of applications. Development efforts have generally focused on achieving desired macroscopic properties via tuning of the IL through variation of the cations and anions. Both the macroscopic and microscopic properties of an IL influence its tunability and thus feasibility of use for selected applications. Works geared toward a microscopic understanding of the nature and strength of the intrinsic cation-anion interactions of ILs have been limited to date. Specifically, the intrinsic strength of the cation-anion interactions in ILs is largely unknown. In previous work, we employed threshold collision-induced dissociation (TCID) approaches supported and enhanced by electronic structure calculations to determine the bond dissociation energies (BDEs) and characterize the nature of the cation-anion interactions in a series of four 2 : 1 clusters of 1-alkyl-3-methylimidazolium cations with the hexafluorophosphate anion, [2Cnmim:PF6]+. To examine the effects of the 1-alkyl chain on the structure and energetics of binding, the cation was varied over the series: 1-ethyl-3-methylimidazolium, [C2mim]+, 1-butyl-3-methylimidazolium, [C4mim]+, 1-hexyl-3-methylimidazolium, [C6mim]+, and 1-octyl-3-methylimidazolium, [C8mim]+. The variation in the strength of binding among these [2Cnmim:PF6]+ clusters was found to be similar in magnitude to the average experimental uncertainty in the measurements. To definitively establish an absolute order of binding among these [2Cnmim:PF6]+ clusters, we extend this work again using TCID and electronic structure theory approaches to include competitive binding studies of three mixed 2 : 1 clusters of 1-alkyl-3-methylimidazolium cations and the hexafluorophosphate anion, [Cn-2mim:PF6:Cnmim]+ for n = 4, 6, and 8. The absolute BDEs of these mixed [Cn-2mim:PF6:Cnmim]+ clusters as well as the absolute difference in the strength of the intrinsic binding interactions as a function of the cation are determined with significantly improved precision. By combining the thermochemical results of the previous independent and present competitive measurements, the BDEs of the [2Cnmim:PF6]+ clusters are both more accurately and more precisely determined. Comparisons are made to results for the analogous [2Cnmim:BF4]+ and [Cn-2mim:BF4:Cnmim]+ clusters previously examined to elucidate the effects of the [PF6]- and [BF4]- anions on the binding.

Structural determination of arginine-linked cisplatin complexes via IRMPD action spectroscopy: arginine binds to platinum via NO- binding mode.

Cisplatin, (NH3)2PtCl2, has been known as a successful metal-based anticancer drug for more than half a century. Its analogue, Argplatin, arginine-linked cisplatin, (Arg)PtCl2, is being investigated because it exhibits reactivity towards DNA and RNA that differs from that of cisplatin. In order to understand the basis for its altered reactivity, the deprotonated and sodium cationized forms of Argplatin, [(Arg-H)PtCl2]- and [(Arg)PtCl2 + Na]+, are examined by infrared multiple photon dissociation (IRMPD) action spectroscopy in the IR fingerprint and hydrogen-stretching regions. Complementary electronic structure calculations are performed using density functional theory approaches to characterize the stable structures of these complexes and to predict their infrared spectra. Comparison of the theoretical IR spectra predicted for various stable conformations of these Argplatin complexes to their measured IRMPD spectra enables determination of the binding mode(s) of Arg to the Pt metal center to be identified. Arginine is found to bind to Pt in a bidentate fashion to the backbone amino nitrogen and carboxylate oxygen atoms in both the [(Arg-H)PtCl2]- and [(Arg)PtCl2 + Na]+ complexes, the NO- binding mode. The neutral side chain of Arg also interacts with the Pt center to achieve additional stabilization in the [(Arg-H)PtCl2]- complex. In contrast, Na+ binds to both chlorido ligands in the [(Arg)PtCl2 + Na]+ complex and the protonated side chain of Arg is stabilized via hydrogen-bonding interactions with the carboxylate moiety. These findings are consistent with condensed-phase results, indicating that the NO- binding mode of arginine to Pt is preserved in the electrospray ionization process even under variable pH and ionic strength.

Influence of 5-Methylation and the 2'- and 3'-Hydroxy Substituents on the Base Pairing Energies of Protonated Cytidine Nucleoside Analogue Base Pairs: Implications for the Stabilities of i-Motif Structures.

Repetitive nucleic acid sequences, which occur in abundance throughout the mammalian genome, are of enormous research interest due to their potential to adopt fascinating and unusual molecular structures such as the i-motif. In remarkable contrast to the DNA double helix, i-motif conformations are stabilized by protonated cytosine base pairs, (Cyt)H+(Cyt), that are centrally located in the core of the i-motif and intercalated vertically in an antiparallel fashion. An in-depth understanding of how modifications influence the stability of i-motif conformations is a prerequisite to understanding their biological functions and the development of effective means of tuning their stability for specific medical and technological applications. Here, the influence of the 2'- and 3'-hydroxy substituents of the sugar moieties and 5-methylation of the cytosine nucleobases on the base-pairing interactions of protonated cytidine nucleoside analogue base pairs, (xCyd)H+(xCyd), are examined by complementary threshold collision-induced dissociation techniques and computational methods. The xCyd nucleosides examined include the canonical DNA and RNA cytidine nucleosides, 2'-deoxycytidine (dCyd) and cytidine (Cyd), as well as several modified cytidine nucleoside analogues, 2',3'-dideoxycytidine (ddCyd), 5-methyl-2'-deoxycytidine (m5dCyd), and 5-methylcytidine (m5Cyd). Comparisons among these model base pairs indicate that the 2'- and 3'-hydroxy substituents of the sugar moieties have very little influence on the strength of the base-pairing interactions, whereas 5-methylation of the cytosine nucleobases is found to enhance the strength of the base-pairing interactions. The increase in stability resulting from 5-methylation is only modest but is more than twice as large for the DNA than RNA protonated cytidine base pair. Overall, present results suggest that canonical DNA i-motif conformations should be more stable than analogous RNA i-motif conformations and that 5-methylation of cytosine residues, a significant epigenetic marker, provides greater stabilization to DNA than RNA i-motif conformations.

Nature and strength of intrinsic cation-anion interactions of 1-alkyl-3-methylimidazolium hexafluorophosphate clusters.

Imidazolium-based cations and the hexafluorophosphate anion are among the most commonly used ionic liquids (ILs). Yet, the nature and strength of the intrinsic cation-anion interactions, and how they influence the macroscopic properties of these ILs are still not well understood. Threshold collision-induced dissociation is utilized to determine the bond dissociation energies (BDEs) of the 2 : 1 clusters of 1-alkyl-3-methylimidazolium cations and the hexafluorophosphate anion, [2Cnmim:PF6]+. The cation, [Cnmim]+, is varied across the series, 1-ethyl-3-methylimidazolium [C2mim]+, 1-butyl-3-methylimidazolium [C4mim]+, 1-hexyl-3-methylimidazolium [C6mim]+, 1-octyl-3-methylimidazolium [C8mim]+, to examine the structural and energetic effects of the size of the 1-alkyl substituent of the cation on the binding to [PF6]-. Complementary electronic structure methods are employed for the [Cnmim]+ cations, (Cnmim:PF6) ion pairs, and [2Cnmim:PF6]+ clusters to elucidate details of the cation-anion interactions and their impact on structure and energetics. Multiple levels of theory are benchmarked with the measured BDEs including B3LYP, B3LYP-GD3BJ, and M06-2X each with the 6-311+G(d,p) basis set for geometry optimizations and frequency analyses and the 6-311+G(2d,2p) basis set for energetic determinations. The modest structural variation among the [Cnmim]+ cations produces only minor structural changes and variation in the measured BDEs of the [2Cnmim:PF6]+ clusters. Present results are compared to those previously reported for the analogous 1-alkyl-3-methylimidazolium tetrafluoroborate IL clusters to compare the effects of these anions on the nature and strength of the intrinsic binding interactions.

Absolute Trends and Accurate and Precise Gas-Phase Binding Energies of 1-Alkyl-3-Methylimidazolium Tetrafluoroborate Ionic Liquid Clusters from Combined Independent and Competitive TCID Measurements.

Ionic liquid (IL) development efforts have focused on achieving desired properties via tuning of the IL through variation of the cations and anions. However, works geared toward a microscopic understanding of the nature and strength of the intrinsic cation-anion interactions of ILs have been rather limited such that the intrinsic strength of the cation-anion interactions in ILs is largely unknown. In previous work, we employed threshold collision-induced dissociation approaches supported and enhanced by electronic structure calculations to characterize the nature of the cation-anion interactions in and determine the bond dissociation energies (BDEs) of a series of four 2:1 clusters of 1-alkyl-3-methylimidazolium cations and tetrafluoroborate anions, [2Cnmim:BF4]+. The cation was varied over the series: 1-ethyl-3-methylimidazolium, [C2mim]+, 1-butyl-3-methylimidazolium, [C4mim]+, 1-hexyl-3-methylimidazolium, [C6mim]+, and 1-octyl-3-methylimidazolium, [C8mim]+, to determine the structural and energetic effects of the size of the 1-alkyl substituent on the binding. The variation in the strength of binding determined for these [2Cnmim:BF4]+ clusters was found to be similar in magnitude to the average experimental uncertainty in these determinations. To definitively establish an absolute order of binding among these [2Cnmim:BF4]+ clusters, we extend this work here to include competitive binding studies of three mixed 2:1 clusters of 1-alkyl-3-methylimidazolium cations and tetrafluoroborate anions, [Cn-2mim:BF4:Cnmim]+ for n = 4, 6, and 8. Importantly, the results of the present work simultaneously provide the absolute BDEs of these mixed [Cn-2mim:BF4:Cnmim]+ clusters and the absolute relative order of the intrinsic binding interactions as a function of the cation with significantly improved precision. Further, by combining the thermochemical results of the previous and present studies, the BDEs of the [2Cnmim:BF4]+ clusters are more accurately and precisely determined.

Gas-Phase Binding Energies and Dissociation Dynamics of 1-Alkyl-3-Methylimidazolium Tetrafluoroborate Ionic Liquid Clusters.

Ionic liquids (ILs) have become increasingly popular due to their useful and unique properties, yet there are still many unanswered questions regarding their fundamental interactions. In particular, details regarding the nature and strength of the intrinsic cation-anion interactions and how they influence the macroscopic properties of ILs are still largely unknown. Elucidating the molecular-level details of these interactions is essential to the development of better models for describing ILs and enabling the purposeful design of ILs with properties tailored for specific applications. Current uses of ILs are widespread and diverse and include applications for energy storage, electrochemistry, designer/green solvents, separations, and space propulsion. To advance the understanding of the energetics, conformations, and dynamics of gas-phase IL clustering relevant to space propulsion, threshold collision-induced dissociation approaches are used to measure the bond dissociation energies (BDEs) of the 2:1 clusters of 1-alkyl-3-methylimidazolium cations and tetrafluoroborate, [2Cnmim:BF4]+. The cation, [Cnmim]+, is varied across the series, 1-ethyl-3-methylimidazolium [C2mim]+, 1-butyl-3-methylimidazolium [C4mim]+, 1-hexyl-3-methylimidazolium [C6mim]+, and 1-octyl-3-methylimidazolium [C8mim]+, to examine the structural and energetic effects of the size of the 1-alkyl substituent on binding. Complementary electronic structure calculations are performed to determine the structures and energetics of the [Cnmim]+ and [BF4]- ions and their binding preferences in the (Cnmim:BF4) ion pairs and [2Cnmim:BF4]+ clusters. Several levels of theory, B3LYP, B3LYP-GD3BJ, and M06-2X, using the 6-311+G(d,p) basis set for geometry optimizations and frequency analyses and the 6-311+G(2d,2p) basis set for energetics, are benchmarked to examine their abilities to properly describe the nature of the binding interactions and to reproduce the measured BDEs. The modest structural variation among these [Cnmim]+ cations produces only minor structural changes and variation in the measured BDEs of the [2Cnmim:BF4]+ clusters. Present findings indicate that the dominant cation-anion interactions involve the 3-methylimidazolium moieties and that these clusters are sufficiently small that differences in packing effects associated with the variable length of the 1-alkyl substituents are not yet significant.

Increased fructose consumption has sex-specific effects on fibroblast growth factor 21 levels in humans.

OBJECTIVE: Fibroblast growth factor 21 (FGF21), a primarily hepatic hormone with pleotropic metabolic effects, is regulated by fructose in humans. Recent work has established that 75 g of oral fructose robustly stimulates FGF21 levels in humans with peak levels occurring 2 h following ingestion; this has been termed an oral fructose tolerance test (OFTT). It is unknown whether prolonged high-fructose consumption influences the FGF21 response to acute fructose or whether biological sex influences FGF21-fructose dynamics. METHODS: Thirty-nine healthy adults underwent baseline OFTT following an overnight fast. For the high-fructose exposure protocol, 20 subjects ingested 75 g of fructose daily for 14 ± 3 d, followed by repeat OFTT. For the control group, an OFTT was repeated following 14 ± 3 d of ad lib diet. For all subjects, FGF21 levels, glucose, insulin, non-esterified fatty acids and triglyceride levels were measured at baseline and 2 h following OFTT. All subjects maintained 3-d food logs prior to OFTT testing. RESULTS: Women demonstrated significantly higher baseline and peak stimulated total and intact FGF21 levels compared with men both before and after high-fructose exposure. Baseline total and intact FGF21 levels decreased following ongoing fructose exposure, maintaining a stable ratio. This decrease was sex specific, with only women demonstrating decreased baseline FGF21 levels. There were no changes in metabolic or anthropometric parameters following the high-fructose exposure. CONCLUSIONS: Daily ingestion of 75 g of fructose for 2 weeks results in a sex-specific decrease in baseline FGF21 levels without change in body weight or biochemical evidence of metabolic injury. There were also sex-specific differences in peak fructose-stimulated FGF21 levels, which do not change with high-fructose consumption. The role of FGF21 in the development of metabolic disease caused by fructose consumption may differ based on biological sex. Future long-term studies should consider sex differences in FGF21-fructose dynamics.

The Management of Stroke Rehabilitation: A Synopsis of the 2019 U.S. Department of Veterans Affairs and U.S. Department of Defense Clinical Practice Guideline.

Description: In June 2019, the U.S. Department of Veterans Affairs (VA) and U.S. Department of Defense (DoD) approved an update of the joint clinical practice guideline for rehabilitation after stroke. This synopsis summarizes the key recommendations from this guideline. Methods: In February 2018, the VA/DoD Evidence-Based Practice Work Group convened a joint VA/DoD guideline development effort that included clinical stakeholders and stroke survivors and conformed to the National Academy of Medicine (formerly the Institute of Medicine) tenets for trustworthy clinical practice guidelines. The guideline panel identified key questions, systematically searched and evaluated the literature, and developed 2 algorithms and 42 key recommendations using the GRADE (Grading of Recommendations Assessment, Development and Evaluation) system. Stroke survivors and their family members were invited to share their perspectives to further inform guideline development. Recommendations: The guideline recommendations provide evidence-based guidance for the rehabilitation care of patients after stroke. The recommendations are applicable to health care providers in both primary care and rehabilitation. Key features of the guideline are recommendations in 6 areas: timing and approach; motor therapy; dysphagia; cognitive, speech, and sensory therapy; mental health therapy; and other functions, such as returning to work and driving.

Structural and Energetic Effects of O2'-Ribose Methylation of Protonated Pyrimidine Nucleosides.

The 2'-substituents distinguish DNA from RNA nucleosides. 2'-O-methylation occurs naturally in RNA and plays important roles in biological processes. Such 2'-modifications may alter the hydrogen-bonding interactions of the nucleoside and thus may affect the conformations of the nucleoside in an RNA chain. Structures of the protonated 2'-O-methylated pyrimidine nucleosides were examined by infrared multiple photon dissociation (IRMPD) action spectroscopy, assisted by electronic structure calculations. The glycosidic bond stabilities of the protonated 2'-O-methylated pyrimidine nucleosides, [Nuom+H]+, were also examined and compared to their DNA and RNA nucleoside analogues via energy-resolved collision-induced dissociation (ER-CID). The preferred sites of protonation of the 2'-O-methylated pyrimidine nucleosides parallel their canonical DNA and RNA nucleoside analogues, [dNuo+H]+ and [Nuo+H]+, yet their nucleobase orientation and sugar puckering differ. The glycosidic bond stabilities of the protonated pyrimidine nucleosides follow the order: [dNuo+H]+ < [Nuo+H]+ < [Nuom+H]+. The slightly altered structures help explain the stabilization induced by 2'-O-methylation of the pyrimidine nucleosides.

Impact of Sodium Cationization on Gas-Phase Conformations of DNA and RNA Cytidine Mononucleotides.

Gas-phase conformations of the sodium-cationized forms of the 2'-deoxycytidine and cytidine mononucleotides, [pdCyd+Na]+ and [pCyd+Na]+, are examined by infrared multiple photon dissociation action spectroscopy. Complimentary electronic structure calculations at the B3LYP/6-311+G(2d,2p)//B3LYP/6-311+G(d,p) level of theory provide candidate conformations and their respective predicted IR spectra for comparison across the IR fingerprint and hydrogen-stretching regions. Comparisons of the predicted IR spectra and the measured infrared multiple photon dissociation action spectra provide insight into the impact of sodium cationization on intrinsic mononucleotide structure. Further, comparison of present results with those reported for the sodium-cationized cytidine nucleoside analogues elucidates the impact of the phosphate moiety on gas-phase structure. Across the neutral, protonated, and sodium-cationized cytidine mononucleotides, a preference for stabilization of the phosphate moiety and nucleobase orientation is observed, although the details of this stabilization differ with the state of cationization. Several low-energy conformations of [pdCyd+Na]+ and [pCyd+Na]+ involving several different orientations of the phosphate moiety and sugar puckering modes are observed experimentally.

Amino acid-linked platinum(II) compounds: non-canonical nucleoside preferences and influence on glycosidic bond stabilities.

Nucleobases serve as ideal targets where drugs bind and exert their anticancer activities. Cisplatin (cisPt) preferentially coordinates to 2'-deoxyguanosine (dGuo) residues within DNA. The dGuo adducts that are formed alter the DNA structure, contributing to inhibition of function and ultimately cancer cell death. Despite its success as an anticancer drug, cisPt has a number of drawbacks that reduce its efficacy, including repair of adducts and drug resistance. Some approaches to overcome this problem involve development of compounds that coordinate to other purine nucleobases, including those found in RNA. In this work, amino acid-linked platinum(II) (AAPt) compounds of alanine and ornithine (AlaPt and OrnPt, respectively) were studied. Their reactivity preferences for DNA and RNA purine nucleosides (i.e., 2'-deoxyadenosine (dAdo), adenosine (Ado), dGuo, and guanosine (Guo)) were determined. The chosen compounds form predominantly monofunctional adducts by reacting at the N1, N3, or N7 positions of purine nucleobases. In addition, features of AAPt compounds that impact the glycosidic bond stability of Ado residues were explored. The glycosidic bond cleavage is activated differentially for AlaPt-Ado and OrnPt-Ado isomers. Formation of unique adducts at non-canonical residues and subsequent destabilization of the glycosidic bonds are important features that could circumvent platinum-based drug resistance.

Infrared multiple photon dissociation action spectroscopy of protonated glycine, histidine, lysine, and arginine complexed with 18-crown-6 ether.

Complexes of 18-crown-6 ether (18C6) with four protonated amino acids (AAs) are examined using infrared multiple photon dissociation (IRMPD) action spectroscopy utilizing light generated by the infrared free electron laser at the Centre Laser Infrarouge d'Orsay (CLIO). The AAs examined in this work include glycine (Gly) and the three basic AAs: histidine (His), lysine (Lys), and arginine (Arg). To identify the (AA)H+(18C6) conformations present in the experimental studies, the measured IRMPD spectra are compared to spectra calculated at the B3LYP/6-311+G(d,p) level of theory. Relative energies of various conformers and isomers are provided by single point energy calculations carried out at the B3LYP, B3P86, M06, and MP2(full) levels using the 6-311+G(2p,2d) basis set. The comparisons between the IRMPD and theoretical IR spectra indicate that 18C6 binds to Gly and His via the protonated backbone amino group, whereas protonated Lys prefers binding via the protonated side-chain amino group. Results for Arg are less definitive with strong evidence for binding to the protonated guanidino side chain (the calculated ground conformer at most levels of theory), but contributions from backbone binding to a zwitterionic structure are likely.

Structures and Relative Glycosidic Bond Stabilities of Protonated 2'-Fluoro-Substituted Purine Nucleosides.

The 2'-substituent is the primary distinguishing feature between DNA and RNA nucleosides. Modifications to this critical position, both naturally occurring and synthetic, can produce biologically valuable nucleoside analogues. The unique properties of fluorine make it particularly interesting and medically useful as a synthetic nucleoside modification. In this work, the effects of 2'-fluoro modification on the protonated gas-phase purine nucleosides are examined using complementary tandem mass spectrometry and computational methods. Direct comparisons are made with previous studies on related nucleosides. Infrared multiple photon dissociation action spectroscopy performed in both the fingerprint and hydrogen-stretching regions allows for the determination of the experimentally populated conformations. The populated conformers of protonated 2'-fluoro-2'-deoxyadenosine, [Adofl+H]+, and 2'-fluoro-2'-deoxyguanosine, [Guofl+H]+, are highly parallel to their respective canonical DNA and RNA counterparts. Both N3 and N1 protonation sites are accessed by [Adofl+H]+, stabilizing syn and anti nucleobase orientations, respectively. N7 protonation and anti nucleobase orientation dominates in [Guofl+H]+. Spectroscopically observable intramolecular hydrogen-bonding interactions with fluorine allow more definitive sugar puckering determinations than possible for the canonical systems. [Adofl+H]+ adopts C2'-endo sugar puckering, whereas [Guofl+H]+ adopts both C2'-endo and C3'-endo sugar puckering. Energy-resolved collision-induced dissociation experiments with survival yield analyses provide relative glycosidic bond stabilities. The N-glycosidic bond stabilities of the protonated 2'-fluoro-substituted purine nucleosides are found to exceed those of their canonical analogues. Further, the N-glycosidic bond stability is found to increase with increasing electronegativity of the 2'-substituent, i.e., H < OH < F. The N-glycosidic bond stability is also greater for the adenine nucleoside analogues than the guanine nucleoside analogues.