Subscribe to RSS
DOI: 10.1055/s-0031-1289672
Synthesis of a Benzophenone C-Nucleoside as Potential Triplet Energy and Charge Donor in Nucleic Acids
Publication History
Publication Date:
23 January 2012 (online)
Abstract
A synthetic route to the C-nucleoside that bears benzophenone as a DNA base substitution directly at the anomeric center of the 2′-deoxyribofuranoside was worked out. Furthermore, the α-anomer of this artificial nucleoside was converted synthetically into the corresponding DNA building block and incorporated into two representative oligonucleotides by automated phosphoramidite chemistry. The chromophore-modified DNA was characterized by methods of optical spectroscopy. The absorption band at ∼350 nm can be used for selective excitation of the benzophenone chromophore outside the nucleic acid absorption range, which makes the benzophenone nucleoside potentially useful for photochemical and photobiological applications.
Key words
DNA - oligonucleotide - phosphorescence - photocatalysis - sensitizer
Benzophenone (Bp) plays a central role in photochemistry and photocatalysis. [¹-8] In the biological context, the photochemistry of Bp and its derivatives is applied extensively for photoaffinity labeling [¹] based on three major advantages: [²] (i) Bp derivatives are chemically more stable than alternative labels (aryl azides and diazirines), especially under conditions of organic synthesis; (ii) the relatively long-lived triplet state of Bp is able to abstract H atoms even from unreactive C-H bonds, such as the α-hydrogen from amino acids [³] in proteins; and (iii) the excitation wavelength of Bp derivatives in the UV-A region does not interfere with the absorption range of proteins (especially tryptophane). On the other hand, in catalytic photochemical reactions, Bp acts as an antenna collecting the light and transferring it to the substrate via sensitization by energy or electron transfer. [4] Both processes have been used for the development of photocatalysts. [5] Template-assisted triplet energy transfer yields enantioselective [2+2] cycloaddition. [6] [7] Photoinduced electron transfer can be applied for enantioselective cyclizations. [8] Both types of C-C bond formation are synthetically highly valuable.
Despite the broad applicability of Bp derivatives, covalent conjugates with nucleic acids can be found very rarely in the literature. [9-¹²] This is surprising since this offers the possibility (i) for new photoaffinity labels to identify DNA- and RNA-binding proteins, and (ii) for new photocatalytically active DNAzymes or ribozymes. Bp-substituted nucleosides and nucleoside analogues have been synthesized as models for ribonucleotide reductases [9] and as photoreactive dyads. [¹0] Bp has been attached to phosphorothioates in RNA [¹¹] and to 2′-deoxyuridine in DNA [¹²] to form interstrand crosslinks. 4-Cyanobenzophenone has been extensively used for electron transfer studies with DNA. [¹³]
Herein, we present the synthesis of a novel C-nucleoside 1 that bears the Bp chromophore as a DNA base substitution directly at the anomeric center of the 2′-deoxyribofuranoside, and its incorporation into oligonucleotides by automated phosphoramidite chemistry. Furthermore, we evaluate preliminarily the influence of the DNA environment on the potential energy and charge transfer properties of 1 by providing the steady-state phosphorescence and phosphorescence lifetimes of the Bp nucleoside in a representative oligonucleotide (DNA1).
The synthesis (Scheme [¹] ) starts from commercially available 4-bromobenzophenone (2), which is converted by standard procedure to the ethylene glycol ketal 3 (87% yield) in order to protect the carbonyl group during the subsequent formation of the nucleosidic bond. On the other hand, Hoffer’s chlorosugar 4 was prepared according to the literature [¹4] to equip the 3′,5′-protected 2′-deoxyribofuranoside moiety with a reactive anomeric center. The Bp 3 was converted into a nucleophile by treatment with magnesium in anhydrous tetrahydrofuran. The subsequent reaction with 4 gave the 3′,5′-protected C-nucleoside 5 in 50% yield, a good value for C-nucleoside preparation. [¹5] The obtained α-selectivity of α/β = 4:1 seems to be typical for this route of C-nucleoside synthesis since it has also been observed by others. [¹5] The anomers, 5α and 5β, were separated by flash chromatography and were assigned based on NOE experiments. The NOE coupling of the conformationally fixed 3′-H was used to differentiate the two 2′-H atoms (Figure [¹] ). The existence of a NOE between either one of the identified 2′-H atoms and the 1′-H elucidates the corresponding anomeric configuration. Careful deprotection of the ketals in the presence of potassium hydrogensulfate [¹6] gives 6α and 6β, respectively. Subsequent treatment with potassium carbonate in anhydrous MeOH cleaves off the two p-toluoyl groups yielding the completely unprotected C-nucleosides 1α and 1β, respectively.
Scheme 1 Synthesis of the benzophenone C-nucleosides 1α/1β, and the DNA building block 8
Due to the better synthetic accessibility, DNA building block 8 was prepared representatively with the α-anomer and applied in automated oligonucleotide synthesis with coupling times of 10 minutes. DNA1 and DNA2 were synthesized as oligonucleotides carrying one or two 1α as DNA substitutions (Figure [¹] ). The sequences of DNA1 and DNA2 are random, but do not bear guanine in the direct vicinity of 1α in order to avoid potentially undesired charge transfer reactions leading to oxidized guanine products. [¹7] [¹8] Adenine was placed representatively as the base opposite to the Bp moiety in DNA1. The melting temperature (Tm) of DNA1 occurs at 56.5 ˚C and reveals a slight destabilization of 4.5 ˚C compared to the corresponding completely unmodified duplex DNA3 (Tm = 61.0 ˚C, with T instead of 1α). This is a typical value for a C-nucleosidic modification. [¹5]
The optical properties of the nucleoside 1α were characterized in comparison with DNA1 and DNA2 by measurements of their absorption in water at room temperature (Figure [²] , top), and phosphorescence at 77 K (Figure [²] , bottom). The UV/Vis absorption spectra of the Bp conjugates reveal the existence of the Bp modification by its characteristic broad side band at ∼350 nm (when compared to the reference DNA3). This band appears to be very small for DNA1, which is not surprising with respect to the ratio between normal DNA bases and the artificial base 1α in the sample. Therefore, DNA2 was synthesized; here the side band is clearly observable due to the double 1α modification. It is important to point out that the absorption band at ∼350 nm can be used for selective excitation outside the nucleic acid absorption range, which is a prerequisite for photochemical and photobiological applications.
Figure 1 NMR and NOE assignments for the H-2′ protons (arrows) and sequences of DNA1 and DNA2; DNA3 is the reference to DNA1 with T instead of the 1α modification
Upon excitation at 355 nm, the steady-state phosphorescence spectrum of the nucleoside 1α in aqueous buffer solution at 77 K shows a similar shape compared to those of the DNA, but reveals slightly altered main maxima. The phosphorescence of 1α has its main maximum at 467 nm, very similar to that of Bp, whereas the phosphorescence maximum of single-stranded DNA1 is blue-shifted and occurs at 440 nm, which shifts further to 444 nm after hybridization with the complementary counterstrand to the duplex (data not shown). Interestingly, a similar blue shift from 467 nm to 442 nm is observed when 1α was dissolved in MeOH instead of aqueous buffer solution. Obviously, the DNA bases behave as a polar organic solvent around 1α in oligonucleotides. This is an important result since it is expected that the photochemical reactivity of the Bp derivative 1α embedded in nucleic acids is more similar to organic solvents (like MeOH) than to water.
Figure 2 Normalized UV absorption spectra at r.t. (top) and normalized phosphorescence at 77 K (bottom) of nucleosides 1α/1β and DNA1-DNA3 (5.0 µM in 10 mM Na-Pi buffer, 250 mM NaCl, pH 7, λexc 355 nm)
The phosphorescence lifetimes of 1α and DNA1 were measured at 77 K and sufficiently fitted by a triexponential (1α) or biexponential (DNA1) decay. Global fit analysis reveals that the overall lifetime of the isolated nucleoside 1α in buffer (2.2 ms) is increased in DNA1 (10.2 ms), which excludes any predominant photoredoxactive process from the triplet state in DNA at 77 K. [¹9-²²] Of course, at room temperature the situation could be different and a full theoretical and physical-chemical account on the singlet and triplet state properties of Bp 1α in DNA (including guanines as adjacent bases) is in preparation. The preliminary result that the overall lifetime of 1α is not shortened, but increased in DNA represents an important feature and advantage of our Bp C-nucleoside and gives the opportunity to develop photocatalytically active DNAzymes based on Bp as a DNA base analogue in aptamers as substrate binding sites. The DNA base environment tunes the phosphorescence properties of the Bp similarly to an organic solvent (MeOH). Using the synthetic Bp C-nucleoside photoaffinity labeling strategies can be envisioned, for example, with DNA-binding proteins and DNA-processing enzymes. In conclusion, C-nucleosides with Bp as an artificial DNA base offer a significant potential to apply this chromophore in nucleic acid related research projects.
Chemicals and anhydrous solvents were purchased and were used without further purification, unless otherwise mentioned. TLC was performed on silica gel coated aluminum foil. Flash chromatography was carried out with silica gel 60. Spectroscopic measurements were recorded in Na-Pi buffer solution (10 mM, pH 7) with 250 mM NaCl using quartz glass cuvettes (10 mm). Absorption spectra were recorded with a spectrometer equipped with a 6 × 6 cell changer unit at 20 ˚C. Mass spectra were measured either in the central analytical facility of the Institute of Organic Chemistry of the University of Regensburg, or at the Institute of Organic Chemistry of KIT, in negative and positive ionization mode. NMR spectra were recorded in deuterated solvents (¹H at 300 MHz, ¹³C at 75 MHz). Chemical shifts are given in ppm relative to TMS. ³¹P NMR was recorded with 85% H3PO4 as external standard.
2-(4-Bromophenyl)-2-phenyl-1,3-dioxolane (3)
In a Dean-Stark apparatus, 4-bromobenzophenone (2; 3.13 g, 12.0 mmol), p-toluenesulfonic acid monohydrate (202 mg, 1.06 mmol), and ethylene glycol (4.23 mL, 75.6 mmol) were dissolved in cyclohexane (20 mL). The reaction mixture was refluxed for 48 h. After cooling to r.t., aq 1 M NaOH (40 mL) was added. The mixture was extracted with Et2O (3 × 30 mL). The combined organic layers were evaporated affording a white solid; yield: 3.18 g (87%); R f = 0.68 (hexanes-EtOAc, 5:1).
¹H NMR (acetone-d 6): δ = 4.04 (s, 4 H, 4-H, 5-H), 7.25-7.38 (m, 3 H, aryl-H), 7.40-7.56 (m, 6 H, aryl-H).
¹³C NMR (acetone-d 6): δ = 65.7 (4-C, 5-C), 109.5 (2-C), 122.4, 126.8, 129.0, 129.1, 132.0, 143.2, 143.2 (aryl-C).
HRMS (ESI): m/z calcd for C15H14BrO2 [M + H]+: 305.0172; found [M + H]+: 305.0356.
1′-[4-(2-Phenyl-1,3-dioxolan-2-yl)phenyl]-3′,5′-di- O - p -toluoyl-1′,2′-didesoxyribose (5)
Activated Mg turnings (134 mg, 5.51 mmol) were stirred in anhyd THF (3 mL) under N2 atmosphere for a few min in a 40 ˚C warm ultrasonic bath. An aliquot (1 mL) of the stock solution of 3 [842 mg, 2.76 mmol in anhyd THF (5 mL)] was added slowly. For further activation, a catalytic amount of MeI can be added. After the reaction mixture had turned red, the remaining amount of 3 (dissolved in THF, 4 mL) was added. This solution was added dropwise to a solution of 4 (670 mg, 1.72 mmol) in anhyd THF (28 mL). The final reaction mixture was stirred at r.t. overnight. Ice water containing NH4Cl (25 mL) was added. The mixture was extracted with EtOAc (4 × 150 mL). The solvent was removed to dryness and the residue was purified by flash chromatography on silica gel (hexanes-EtOAc 12:1) to obtain a white solid as an anomeric mixture α/β = 4:1; yield: 498 mg (50%). Separation of the two anomers can be achieved by column chromatography on silica gel (hexanes-EtOAc, 12:1); R f = 0.26 (α), 0.31 (β) (hexanes-EtOAc, 5:1).
¹H NMR (CDCl3): δ (α-anomer) = 2.27-2.34 (m, 1 H, 2′-H), 2.39 (s, 3 H, ArCH 3), 2.41 (s, 3 H, ArCH 3), 2.59-2.96 (m, 1 H, 2′-H), 4.00-4.10 (m, 4 H, dioxolane-H), 4.53-4.62 (m, 2 H, 5′-H), 4.66-4.70 (m, 1 H, 4′-H), 5.36 (t, J = 6.7 Hz, 1 H, 1′-H), 5.58-5.62 (m, 1 H, 3′-H), 7.12 (d, J = 8.1 Hz, 2 H, p-Tol-5′-H,), 7.24 (d, J = 7.9 Hz, 2 H, p-Tol-3′-H), 7.27-7.34 (m, 3 H, aryl-H), 7.40 (d, J = 8.2 Hz, 2 H, aryl-H), 7.50-7.54 (m, 4 H, aryl-H), 7.69 (d, J = 8.1 Hz, 2 H, p-Tol-aryl-H), 7.96 (d, J = 8.1 Hz, 2 H, p-Tol-aryl-H); δ (β-anomer) = 2.15-2.28 (m, 1 H, 2′-H), 2.40 (s, 3 H, aryl-CH 3), 2.43 (s, 3 H, aryl-CH 3), 2.46-2.55 (m, 1 H, 2′-H), 4.02-4.08 (m, 4 H, dioxolane-H), 4.49-4.56 (m, 1 H, 4′-H), 4.59-4.66 (m, 2 H, 5′-H), 5.23 (dd, J = 5.0, 10.9 Hz, 1 H, 1′-H), 5.55-5.63 (m, 1 H, 3′-H), 7.20 (d, J = 8.0 Hz, 2 H, p-Tol-5′-H), 7.27-7.40 (m, 7 H, aryl-H), 7.46-7.53 (m, 4 H, aryl-H), 7.87-8.02 (m, 4 H, aryl-H).
¹³C NMR (CDCl3): δ (α-anomer) = 21.8, 21.8 (aryl-CH3), 40.4 (2′-C), 64.7 (5′-C), 64.9, 65.0 (dioxolane-C), 76.5 (3′-C), 80.2 (1′-C), 82.3 (4′-C), 109.5 (dioxolane-C), 125.6, 126.3, 126.3, 126.4 (aryl-C), 126.9, 127.2 (p-Tol-C), 128.0, 128.0, 128.3 (aryl-C), 129.2, 129.3, 129.8, 129.9 (p-Tol-aryl-C), 141.5, 142.2, 142.3 (aryl-C), 144.0, 144.0, 166.3, 166.6 (p-Tol-C); δ (β-anomer) = 21.8, 21.9 (aryl-CH3), 41.8 (2′-C), 65.0 (5′-C), 64.9, 65.0 (dioxolane-C), 77.4 (3′-C), 80.8 (1′-C), 83.1 (4′-C), 109.4 (dioxolane-C), 125.9, 126.3, 126.5 (aryl-C), 127.1, 127.2 (p-Tol-C), 128.2, 128.3 (aryl-C), 129.3, 129.3, 129.8, 129.9 (p-Tol-aryl-C), 140.6, 142.0, 142.1 (aryl-C), 143.9, 144.3, 166.3, 166.5 (p-Tol-C).
HRMS (ESI): m/z calcd for C36H35O7 [M + H]+: 579.2377; found [M + H]+: 579.2419.
1′-(4-Benzoylphenyl)-3′,5′-di- O - p -toluoyl-1′,2′-didesoxyribose (6)
Compound 5 (32 mg, 0.06 mmol), KHSO4 (82 mg), and wet silica gel (117 mg, 60% w/w silica gel + 40% w/w H2O) were added to CH2Cl2 (2 mL). This suspension was refluxed for 1 h. The silica gel was filtered off and washed with EtOAc (10 mL). The organic layer was evaporated affording a slightly yellow solid; yield: 29 mg (98%); R f = 0.33 (α), 0.38 (β) (toluene-EtOAc, 15:1).
¹H NMR (CDCl3): δ (α-anomer) = 2.31-2.46 (m, 7 H, 2′ + aryl-CH 3), 2.91-3.05 (m, 1 H, 2′-H), 4.55-4.64 (m, 2 H, 5′-H), 4.70-4.77 (m, 1 H, 4′-H), 5.48 (dd, J = 4.9, 7.7 Hz, 1 H, 1′-H), 5.57-5.66 (m, 1 H, 3′-H), 7.11-8.01 (m, 17 H, aryl-H); δ (β-anomer) = 2.18-2.31 (m, 1 H, 2′-H), 2.39 (s, 3 H, aryl-CH 3), 2.44 (s, 3 H, aryl-CH 3), 2.56-2.66 (m, 1 H, 2′-H), 4.55-4.61 (m, 1 H, 4′-H), 4.61-4.74 (m, 2 H, 5′-H), 5.34 (dd, J = 4.9, 10.7 Hz, 1 H, 1′-H), 5.60-5.66 (m, 1 H, 3′-H), 7.15-8.02 (m, 17 H, aryl-H).
¹³C NMR (CDCl3): δ (α-anomer) = 21.8, 21.9 (aryl-CH3), 40.5 (2′-C), 64.7 (5′-C), 76.5 (3′-C), 80.0 (1′-C), 82.7 (4′-C), 125.5, 126.8 (aryl-C), 127.2 (p-Tol-C), 128.4, 128.4 (aryl-C), 129.2, 129.2, 129.3, 129.7, 129.89, 130.2, 130.5, 132.6, 136.7, 137.8, 144.1, 144.3 (p-Tol-aryl-C), 147.7 (aryl-C), 166.1, 166.5 (p-Tol-C), 196.5 (C=O); δ = (β-anomer) = 21.8, 21.9 (aryl-CH3), 41.9 (2′-C), 64.8 (5′-C), 77.4 (3′-C), 80.5 (1′-C), 83.4 (4′-C), 125.8, 128.4 (aryl-C), 129.4, 129.4, 129.8, 129.9, 130.2, 130.5, 131.4, 132.6, 135.1, 137.2, 144.1, 144.4 (p-Tol-aryl-C), 145.7 (aryl-C), 166.3, 166.5 (p-Tol-C), 196.5 (C=O).
HRMS (ESI): m/z calcd for C34H31O6 [M + H]+: 535.2115; found [M + H]+: 535.2142.
1′-(4-Benzoylphenyl)-1′,2′-didesoxyribose (1α)
Compound 6α (157 mg, 0.29 mmol) was dissolved in anhyd MeOH (5.5 mL), and K2CO3 (89 mg) was added. The reaction mixture was stirred at r.t. overnight until K2CO3 was completely dissolved. The solvent was removed under reduced pressure and the residue was purified by flash chromatography on silica gel (CH2Cl2-MeOH, 15:1) to afford a white solid; yield: 62 mg (71%); R f = 0.29 (α), 0.37 (β) (CH2Cl2-MeOH, 9:1).
¹H NMR (MeOD): δ = 1.89-1.96 (m, 1 H, 2′-H), 2.70-2.77 (m, 1 H, 2′-H), 3.64-3.79 (m, 2 H, 5′-H), 4.01-4.05 (m, 1 H, 4′-H), 4.37-4.42 (m, 1 H, 3′-H), 5.17 (t, J = 7.7 Hz, 1 H, 1′-H), 7.50-7.55 (m, 2 H, aryl-H), 7.55-7.60 (m, 2 H, aryl-H), 7.61-7.66 (m, 1 H, aryl-H), 7.74-7.79 (m, 4 H, aryl-H).
¹³C NMR (MeOD): δ = 44.7 (2′-C), 63.3 (5′-C), 73.5 (3′-C), 80.4 (1′-C), 87.7 (4′-C), 126.8, 129.5, 131.0, 131.3, 133.7, 137.7, 139.0, 150.2 (aryl-C), 198.4 (C=O).
HRMS (ESI): m/z calcd for C18H19O4 [M + H]+: 299.1278; found [M + H]+: 299.1305.
1′-(4-Benzoylphenyl)-5′- O -(dimethoxytrityl)-1′,2′-didesoxyribose (7)
Compound 1α (214 mg, 0.72 mmol) was dissolved in anhyd pyridine (17.8 mL) under N2 atmosphere. 4,4′-Dimethoxytriphenylmethyl chloride (267 mg, 0.79 mmol) was added and the reaction mixture was stirred at r.t. for 45 h. The solvent was removed under reduced pressure and the residue was purified by flash chromatography on silica gel [hexanes-EtOAc (4:1) + 0.01% Et3N] to give a slightly yellow solid; yield: 311 mg (72%); R f = 0.15 (hexanes-EtOAc, 3:1).
¹H NMR (MeOD): δ = 1.89-1.96 (m, 1 H, 2′-H), 2.71-2.79 (m, 1 H, 2′-H), 3.21-3.25 (m, 1 H, 5′-H), 3.28-3.34 (m, 1 H, 5′-H), 3.78 (s, 6 H, OCH3), 4.16-4.21 (m, 1 H, 4′-H), 4.41-4.46 (m, 1 H, 3′-H), 5.23 (t, J = 7.6 Hz, 1 H, 1′-H), 6.83-6.89 (m, 4 H, aryl-H), 7.17-7.81 (m, 18 H, aryl-H).
¹³C NMR (MeOD): δ = 44.7 (2′-C), 55.7 (OCH3), 65.5 (5′-C), 74.4 (3′-C), 80.8 (1′-C), 86.9 (4′-C), 114.0, 114.1, 127.0, 128.7, 128.8, 129.3, 129.4, 129.5, 131.0, 131.3, 131.3, 131.4, 133.8, 137.4, 137.4, 137.4, 137.8, 139.0, 146.6, 150.0, 160.1, 160.1 (aryl-C), 198.4 (C=O).
HRMS (ESI): m/z calcd for C39H36O6 + K [M + K]+: 639.2143; found [M + K]+: 639.2177.
1′-(4-Benzoylphenyl)-3′- O -(2-cyanoethoxydiisopropylaminophosphanyl)-5′- O -(dimethoxytrityl)-1′,2′-didesoxyribose (8)
Compound 7 (45 mg, 0.07 mmol) was dissolved in anhyd CH2Cl2 (4.5 mL) under N2 atmosphere. i-Pr2NEt (45 µL, 0.26 mmol) was added and the reaction mixture was stirred at r.t. for 10 min. 2-Cyanoethoxydiisopropylaminophosphanyl chloride (25 µL, 0.11 mmol) was added and the solution was stirred at r.t. for 2 h. More 2-cyanoethoxydiisopropylaminophosphanyl chloride (12 µL, 0.05 mmol) was added and the mixture was stirred at r.t. for another 2 h. The mixture was directly transferred on a silica gel column for purification [hexanes-EtOAc (2:1) + 0.1% Et3N]. Flash chromatography afforded a white foam; yield: 39 mg (65%); R f = 0.38, 0.44 (hexanes-EtOAc, 3:1).
¹H NMR (DMSO-d 6): δ = 0.92-1.00 (m, 6 H, i-Pr-CH 3), 1.05-1.10 [m, 6 H, CH(CH 3)2], 1.92-2.09 (m, 1 H, 2′-H), 2.56-2.66 (m, 2 H, CH2), 2.73-2.81 (m, 1 H, 2′-H), 3.06-3.12 (m, 1 H, 5′-H), 3.16-3.24 (m, 1 H, 5′-H), 3.37-3.48 (m, 2 H, CH2), 3.51-3.58 [m, 2 H, CH(CH3)2], 3.71-3.76 (m, 6 H, OCH3), 4.21-4.31 (m, 1 H, 4′-H), 4.44-4.53 (m, 1 H, 3′-H), 5.25-5.34 (m, 1 H, 1′-H), 6.87-6.93 (m, 4 H, aryl-H), 7.20-7.77 (m, 18 H, aryl-H).
¹³C NMR (DMSO-d 6): δ = 19.7, 19.7 (CH2), 24.1, 24.2, 24.2, 24.3, 24.3, 24.3 [CH(CH3)2], 41.9 (2′-C), 42.4, 42.5 (CH2), 55.0 (OCH3), 58.0, 58.1, 58.2 [CH(CH3)2], 63.8 (5′-C), 74.59, 75.0 (3′-C), 78.8, 78.9 (1′-C), 85.5 (4′-C), 113.2 (aryl-C), 118.9, 118.7 (CN), 125.7, 125.8, 126.7, 127.7, 127.7, 127.8, 128.6, 129.5, 129.5, 129.7, 129.7, 129.7, 129.8, 132.6, 135.6, 135.8, 137.2, 144.9, 158.1 (aryl-C), 195.46 (C=O).
³¹P NMR (DMSO-d 6): δ = 148.21, 148.45.
MS (ESI): m/z calcd for C48H54N2O7P [M + H]+: 801.4; found [M + H]+: 801.4.
Oligonucleotides
Unmodified oligonucleotides were purchased and used without further purification. Bp-modified DNA1 and DNA2 were prepared using standard phosphoramidite chemistry. Reagents and controlled pore glass (CPG) (1 µmol) were purchased and used without further purification. The concentration of 8 was increased to 0.1 M (MeCN). After preparation, the oligonucleotides were cleaved from the resin and deprotected by treatment with concd NH4OH at 50 ˚C for 24 h. The oligonucleotides were purified by semi-preparative HPLC column (RP-18), using the following gradient, 0-15% B over 50 min (A: 50 mM NH4OAc buffer, B: MeCN), and identified by MS (ESI).
DNA1
M = 5222.9.
MS (ESI): m/z calcd for (C174H210N60O99P16)4- [M - 4 H]4-: 1304.7; (C174H211N60O99P16)³- [M - 3 H]³-: 1740.0; found [M - 4 H]4-: 1305.1, [M - 3 H]³-: 1740.5.
DNA2
M = 5233.0.
MS (ESI): m/z calcd for (C180H212N60O95P16)4- [M - 4 H]4-: 1307.2; (C180H213N60O95P16)³- [M - 3 H]³-: 1743.3.0; found [M - 4 H]4-: 1307.7, [M - 3 H]³-: 1744.0.
Supporting Information for this article is available online at http://www.thieme-connect.com.accesdistant.sorbonne-universite.fr/ejournals/toc/synthesis. Included are images of ¹H NMR, ¹³C NMR, ³¹P NMR and mass spectra of the synthesized nucleotides, as well as images of HPLC traces and mass and phosphorescence lifetime spectra of 1α and DNA1.
Acknowledgment
Financial support by GRK 1626 (Chemical Photocatalysis) and KIT is gratefully acknowledged. We thank Hartmut Yersin and his group (Chemistry Department, University of Regensburg, Germany) for measurement of the phosphorescence spectra and lifetimes.
- 1 See review:
Fleming SA. Tetrahedron 1995, 51: 12479 - 2 See review:
Dormán G.Prestwich GD. Biochemistry 1994, 33: 5661 - 3
Galardy RE.Craig LC.Jamieson JD.Printz MP. J. Biol. Chem. 1974, 249: 3510 - See reviews:
- 4a
Fagnoni M.Dondi D.Ravelli D.Albini A. Chem. Rev. 2007, 107: 2725 - 4b
Ravelli D.Dondi D.Fagnoni M.Albini A. Chem. Soc. Rev. 2009, 38: 1999 - See reviews:
- 5a
Svoboda J.König B. Chem. Rev. 2006, 106: 5413 - 5b
Wessig P. Angew. Chem. Int. Ed. 2006, 45: 2168 - 6a
Müller C.Bauer A.Bach T. Angew. Chem. Int. Ed. 2009, 48: 6640 - 6b
Müller C.Bauer A.Maturi MM.Cuquerella MC.Miranda MA.Bach T. J. Am. Chem. Soc. 2011, 133: 16689 - 7
Kauble DF.Lynch V.Krische MJ. J. Org. Chem. 2003, 68: 15 - 8
Bauer A.Westkämper F.Grimme S.Bach T. Nature 2005, 436: 1139 - 9a
Lehmann TE.Berkessel A. J. Org. Chem. 1997, 62: 302 - 9b
Lehmann TE.Müller G.Berkessel A. J. Org. Chem. 2000, 65: 2508 - 10
Paris C.Encinas S.Belmadoui N.Climent MJ.Miranda MA. Org. Lett. 2008, 10: 4409 - 11
Musier-Forsyth K.Schimmel P. Biochemistry 1994, 33: 773 - 12
Nakatani K.Yoshida T.Saito I. J. Am. Chem. Soc. 2002, 124: 2118 - 13
Nakatani K.Saito I. Top. Curr. Chem. 2004, 236: 163 - 14a
Hoffer M. Chem. Ber. 1960, 93: 2777 - 14b
Rolland V.Kotera M.Lhomme J. Synth. Commun. 1997, 27: 3505 - 14c
Dhimitruka I.SantaLucia J. Synlett 2004, 335 - 14d
Chin T.-M.Huang L.-K.Kan L.-S. J. Chin. Chem. Soc. 1997, 44: 413 - 15
Štambaský J.Hocek M.Kočovský P. Chem. Rev. 2009, 109: 6729 - 16
Mirjalili BF.Zolfigol MA.Bamoniri A.Hazar A. Bull. Korean Chem. Soc. 2004, 25: 1075 - 17
Adam W.Arnold MA.Nau WM.Pischel U.Saha-Möller CR. J. Am. Chem. Soc. 2002, 124: 3893 - 18a
Douki T.Cadet J. Int. J. Radiat. Biol. 1999, 75: 571 - 18b
Morin B.Cadet J. Photochem. Photobiol. 1994, 60: 102 - 19
Delatour T.Douki T.D’Ham C.Cadet J. Photochem. Photobiol. 1998, 44: 191 - 20
Hélène C.Charlier M. Biochimie 1971, 53: 1175 - 21
Lamola AA. J. Am. Chem. Soc. 1966, 88: 813 - 22
Jennings BH.Pastra S.-C.Wellington JL. Photochem. Photobiol. 1970, 11: 215
References
- 1 See review:
Fleming SA. Tetrahedron 1995, 51: 12479 - 2 See review:
Dormán G.Prestwich GD. Biochemistry 1994, 33: 5661 - 3
Galardy RE.Craig LC.Jamieson JD.Printz MP. J. Biol. Chem. 1974, 249: 3510 - See reviews:
- 4a
Fagnoni M.Dondi D.Ravelli D.Albini A. Chem. Rev. 2007, 107: 2725 - 4b
Ravelli D.Dondi D.Fagnoni M.Albini A. Chem. Soc. Rev. 2009, 38: 1999 - See reviews:
- 5a
Svoboda J.König B. Chem. Rev. 2006, 106: 5413 - 5b
Wessig P. Angew. Chem. Int. Ed. 2006, 45: 2168 - 6a
Müller C.Bauer A.Bach T. Angew. Chem. Int. Ed. 2009, 48: 6640 - 6b
Müller C.Bauer A.Maturi MM.Cuquerella MC.Miranda MA.Bach T. J. Am. Chem. Soc. 2011, 133: 16689 - 7
Kauble DF.Lynch V.Krische MJ. J. Org. Chem. 2003, 68: 15 - 8
Bauer A.Westkämper F.Grimme S.Bach T. Nature 2005, 436: 1139 - 9a
Lehmann TE.Berkessel A. J. Org. Chem. 1997, 62: 302 - 9b
Lehmann TE.Müller G.Berkessel A. J. Org. Chem. 2000, 65: 2508 - 10
Paris C.Encinas S.Belmadoui N.Climent MJ.Miranda MA. Org. Lett. 2008, 10: 4409 - 11
Musier-Forsyth K.Schimmel P. Biochemistry 1994, 33: 773 - 12
Nakatani K.Yoshida T.Saito I. J. Am. Chem. Soc. 2002, 124: 2118 - 13
Nakatani K.Saito I. Top. Curr. Chem. 2004, 236: 163 - 14a
Hoffer M. Chem. Ber. 1960, 93: 2777 - 14b
Rolland V.Kotera M.Lhomme J. Synth. Commun. 1997, 27: 3505 - 14c
Dhimitruka I.SantaLucia J. Synlett 2004, 335 - 14d
Chin T.-M.Huang L.-K.Kan L.-S. J. Chin. Chem. Soc. 1997, 44: 413 - 15
Štambaský J.Hocek M.Kočovský P. Chem. Rev. 2009, 109: 6729 - 16
Mirjalili BF.Zolfigol MA.Bamoniri A.Hazar A. Bull. Korean Chem. Soc. 2004, 25: 1075 - 17
Adam W.Arnold MA.Nau WM.Pischel U.Saha-Möller CR. J. Am. Chem. Soc. 2002, 124: 3893 - 18a
Douki T.Cadet J. Int. J. Radiat. Biol. 1999, 75: 571 - 18b
Morin B.Cadet J. Photochem. Photobiol. 1994, 60: 102 - 19
Delatour T.Douki T.D’Ham C.Cadet J. Photochem. Photobiol. 1998, 44: 191 - 20
Hélène C.Charlier M. Biochimie 1971, 53: 1175 - 21
Lamola AA. J. Am. Chem. Soc. 1966, 88: 813 - 22
Jennings BH.Pastra S.-C.Wellington JL. Photochem. Photobiol. 1970, 11: 215
References
Scheme 1 Synthesis of the benzophenone C-nucleosides 1α/1β, and the DNA building block 8
Figure 1 NMR and NOE assignments for the H-2′ protons (arrows) and sequences of DNA1 and DNA2; DNA3 is the reference to DNA1 with T instead of the 1α modification
Figure 2 Normalized UV absorption spectra at r.t. (top) and normalized phosphorescence at 77 K (bottom) of nucleosides 1α/1β and DNA1-DNA3 (5.0 µM in 10 mM Na-Pi buffer, 250 mM NaCl, pH 7, λexc 355 nm)