Fluorescence enhancement of oligodeoxynucleotides modified with green fluorescent protein chromophore mimics upon triplex formation

Kanamori, Takashi; Takamura, Akihiro; Tago, Nobuhiro; Yoshio, Masaki; Ohkubo, Akihiro; Sekine, Mitsuo; Seio, Kohji

doi:10.1039/c6ob01278g

Cited by 17 publications

(4 citation statements)

References 38 publications

(16 reference statements)

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“…Fluorescent labeling is important in diagnostics and imaging of nucleic acids. − Diverse environmentally sensitive fluorophores can be attached to nucleobases for applications in sensing changes in the microenvironment including changes of secondary structures, − detection of hybridization, − mismatches, or interactions with proteins. − Several solvatochromic DNA labels were reported − to change the emission wavelength by changing the polarity of the environment, and typically an interaction of DNA with protein resulted in a blue shift and increased the fluorescence intensity. On the other hand, fluorophores based on molecular rotors are viscosity-sensitive and can be used to detect protein–DNA interactions.…”

Section: Introductionmentioning

confidence: 99%

Nucleotides Bearing Red Viscosity-Sensitive Dimethoxy-Bodipy Fluorophore for Enzymatic Incorporation and DNA Labeling

et al. 2022

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Nucleosides and 2′-deoxyribonucleoside triphosphates (dNTPs) bearing 3,3′-dimethoxy-2,2′-diphenyl-6-(4-hydroxyphenyl)-bodipy fluorophore attached through a propargyl or propargyl-triethylene glycol linker to position 5 of 2′-deoxycytidine were designed and synthesized. They exerted bright red fluorescence and good sensitivity to viscosity changing their lifetime from 1.6 to 4.5 ns. The modifed dNTPs were substrates for DNA polymerases and were used in enzymatic synthesis of labeled DNA through primer extension. The modified DNA probes served as viscosity sensors responding to protein binding by changes of lifetime. The nucleotide with longer linker (dC pegMOB TP) was transported to live cells and incorporated into the genomic DNA, which can be useful for staining of DNA and imaging of DNA synthesis.

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Section: Introductionmentioning

confidence: 99%

Nucleotides Bearing Red Viscosity-Sensitive Dimethoxy-Bodipy Fluorophore for Enzymatic Incorporation and DNA Labeling

et al. 2022

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“…HBI analogs, particularly, 3,5-difluoro-4-hydroxybenzylidene imidazolinone (DFHBI), 3 (Figure ), were reported to exhibit improved photophysical properties in comparison to HBI. , The enhancement of the photophysical properties of DFHBI vs HBI is due to the stabilization of the phenolate form by the fluorine atoms. − …”

Section: Introductionmentioning

confidence: 99%

“…In addition, DFHBI has been reported for improving photophysical properties of HBI, causing red shift of the absorption and emission wavelengths . Recently, new switchable fluorescent nucleosides, 5-[( Z )-4-(4-hydroxybenzylidene)-2-methyl-5-oxo-imidazolin-1-yl]-2′-deoxyuridine, 4a , and 5-[( Z )-4-(3,5-difluoro-4-hydroxybenzylidene)-2-methyl-5-oxoimidazolin-1-yl]-2′-deoxyuridine, 4b (Figure ), were reported as well as their incorporation into oligonucleotides, resulting in the enhancement of photophysical properties for oligonucleotides incorporating 4b …”

Section: Introductionmentioning

confidence: 99%

“…14 Recently, new switchable fluorescent nucleosides, 5-[(Z)-4-(4-hydroxybenzylidene)-2-methyl-5-oxoimidazolin-1-yl]-2′-deoxyuridine, 4a, and 5-[(Z)-4-(3,5-difluoro-4-hydroxybenzylidene)-2-methyl-5-oxoimidazolin-1-yl]-2′-deoxyuridine, 4b (Figure 4), were reported as well as their incorporation into oligonucleotides, resulting in the enhancement of photophysical properties for oligonucleotides incorporating 4b. 11 In this study, we targeted the development of new fluorescent probes with enhanced signal to background ratio, and improved properties as compared to dU HBI −oligonucleotides, 15 as well as the quantification of target mRNA in the sample, and determination of the mode of binding of the fluorescent moiety. Specifically, we report on the synthesis of a NIC monomer dU DFHBI , 5a (Figure 5), and its photophysical properties compared to dU HBI .…”

Section: ■ Introductionmentioning

confidence: 99%

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Specific, Sensitive, and Quantitative Detection of HER-2 mRNA Breast Cancer Marker by Fluorescent Light-Up Hybridization Probes

et al. 2020

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Currently, there is demand for fluorescent oligonucleotide probes for diagnostic purposes. To address this necessity, we developed nucleosides containing a flexible spacer with an intercalating moiety at its end (NIC molecules). The intercalator is based on 4-hydroxybenzylidene imidazolinone (HBI), found in the Green Fluorescent Protein. We synthesized 20-mer oligonucleotides, ON1−ON4, incorporating the DMTr phosphorodiamidite monomer of dU HBI , 2, and the corresponding dU DFHBI , 5b, monomer. ON1−ON4 target the HER-2 mRNA breast cancer marker for the diagnostics of breast cancer subtype. Hybridization of ON1/ON2 and ON3/ON4 with complementary 2′-OMe-RNA resulted in emission at 462 and 481 nm, respectively, and up to 46-fold increase in fluorescence intensity. CD and 19 F-NMR data indicated that HBI and DFHBI fluorophores bind as intercalators and stabilize the duplexes (up to ΔT m 6 °C). Furthermore, addition of ON1−ON4 to total RNA extracted from cancer cells that overexpress HER-2 mRNA, resulted in a significant fluorescence enhancement of ON3 and ON4. The latter sensitively detected low concentrations of the target mRNA (at total RNA 30 ng/μL). These probes were photostable for 200 min. Using a dilution curve, we quantified the number of HER-2 transcripts in a cell. In conclusion, ON3 and ON4 are promising diagnostic probes for an easy, instantaneous, specific, and sensitive detection of levels of oncogenes. Importantly, the NIC concept, demonstrated here for diagnostics of breast cancer, is universal and may be applied not only in a clinical setting but also for the detection of any RNA.

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Nucleotide‐Bearing Benzylidene‐Tetrahydroxanthylium Near‐IR Fluorophore for Sensing DNA Replication, Secondary Structures and Interactions

Kuba

Kraus

Pohl

et al. 2020

Chemistry A European J

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Thymidine triphosphate bearing benzylidene‐tetrahydroxanthylium near‐IR fluorophore linked to the 5‐methyl group via triazole was synthesized through the CuAAC reaction and was used for polymerase synthesis of labelled DNA probes. The fluorophore lights up upon incorporation to DNA (up to 348‐times) presumably due to interactions in major groove and the fluorescence further increases in the single‐stranded oligonucleotide. The labelled dsDNA senses binding of small molecules and proteins by a strong decrease of fluorescence. The nucleotide was used as a light‐up building block in real‐time PCR for detection of SARS‐CoV‐2 virus.

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Fluorescence enhancement of oligodeoxynucleotides modified with green fluorescent protein chromophore mimics upon triplex formation

Cited by 17 publications

References 38 publications

Nucleotides Bearing Red Viscosity-Sensitive Dimethoxy-Bodipy Fluorophore for Enzymatic Incorporation and DNA Labeling

Nucleotides Bearing Red Viscosity-Sensitive Dimethoxy-Bodipy Fluorophore for Enzymatic Incorporation and DNA Labeling

Specific, Sensitive, and Quantitative Detection of HER-2 mRNA Breast Cancer Marker by Fluorescent Light-Up Hybridization Probes

Nucleotide‐Bearing Benzylidene‐Tetrahydroxanthylium Near‐IR Fluorophore for Sensing DNA Replication, Secondary Structures and Interactions

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