Photolabile Precursors of Cyclic Nucleotides with High Aqueous Solubility and Stability

Wang, Limin; Corrie, John E. T.; Wootton, John F.

doi:10.1021/jo020040g

Cited by 23 publications

(22 citation statements)

References 28 publications

(25 reference statements)

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“…Improvements in one or more of their properties (solubility, stability, rate of uncaging and two-photon cross-section) have been reported, however. For example, NPE-cAMP is not very soluble at pH 7, so a highly soluble version of this compound has been synthesized in nine synthetic steps 34 , which has the added benefit of also being more hydrolytically stable. The rate of uncaging of NPE-ATP is a little slow for some studies (rate is 83 s |[minus]|1 ), so many faster-uncaging caged ATP probes have been synthesized.…”

Section: Caged Nucleotides Nucleosides and Inositolsmentioning

confidence: 99%

“…Some substrates are so highly soluble (for example, EGTA, IP 3 ) that adding the caging chromophore to these molecules makes little difference to their solubility, whereas others can be profoundly affected by caging (for example, serotonin, cAMP, GABA, DAG, sphingosine-1-phosphate). This problem is often ignored in the literature, but can be solved by adding additional (usually negative) charge to the caging chromophore, which unfortunately complicates the synthesis considerably 34 .…”

Section: Chemical Considerationsmentioning

confidence: 99%

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Caged compounds: photorelease technology for control of cellular chemistry and physiology

2007

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Caged compounds are light-sensitive probes that functionally encapsulate biomolecules in an inactive form. Irradiation liberates the trapped molecule, permitting targeted perturbation of a biological process. Uncaging technology and fluorescence microscopy are 'optically orthogonal': the former allows control, and the latter, observation of cellular function. Used in conjunction with other technologies (for example, patch clamp and/or genetics), the light beam becomes a uniquely powerful tool to stimulate a selected biological target in space or time. Here I describe important examples of widely used caged compounds, their design features and synthesis, as well as practical details of how to use them with living cells.The idea behind the caging technique is that a molecule of interest can be rendered biologically inert (or caged) by chemical modification with a photoremovable protecting group (Fig. 1). Illumination results in a concentration jump of the biologically active molecule that can bind to its cellular receptor, switching on (or off) the targeted process. Virtually every kind of signaling molecule or second messenger, of every size| [mdash]|from protons to proteins| [mdash]|has been caged 1 .Why are caged compounds so useful? A single component of cellular chemistry can control the function of a cell, and such cellular regulation can be temporally or spatially defined, intracellular or extracellular, and amplitude-or frequency-modulated 2 . Photomanipulation of cellular chemistry using caged compounds provides a uniquely powerful means to interact with such cellular dynamics, as it can touch upon any one of the above dimensions. Thus, since light passes through cell membranes, uncaging can rapidly release a biomolecule in an intracellular compartment. This space is not readily accessible to many second messengers (for example, inositol-1,4,5-trisphosphate (IP 3 ), ATP, Ca 2+ , cAMP, cGMP) when they are applied to cells externally, as their charge makes them impermeable to the plasma membrane. Furthermore, uniform illumination results in release throughout the cytosol, or the release can be localized by focusing the uncaging beam on one part of a cell. Likewise, extracellular uncaging of neurotransmitters and hormones is tunable, allowing stimulation of many neurons simultaneously or of single synapses| [mdash]|by global or focused illumination, respectively. Light cannot only be directed, but also modulated in time andCorrespondence should be addressed to Graham C R Ellis-Davies (cagedca@hotmail.com). (Fig. 2) has been widely used to study many Ca 2+ -controlled processes. In particular, secretory processes in neuronal and non-neuronal cells have been extensively studied with caged calcium 22 . Rapid uncaging of glutamate using two-photon photolysis is particularly useful for the rational stimulation of visually identified synapses in complex tissue preparations such as acutely isolated brain slices from the hippocampus 23 (Fig. 2b). Finally, uncaging of mRNA in vivo in zebrafish is an excellen...

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Section: Caged Nucleotides Nucleosides and Inositolsmentioning

confidence: 99%

Section: Chemical Considerationsmentioning

confidence: 99%

Caged compounds: photorelease technology for control of cellular chemistry and physiology

2007

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“…Flash column chromatography was carried out on 43±60 mesh silica gel. NMR spectra were recorded (JEOL GSX270) at 270 MHz for 1 H and at 67 MHz for 13 C with CDCl 3 as a solvent and TMS as an internal standard unless otherwise specified. IR spectra were recorded on a Thermo Nicolet Avatar 320 instrument in ATR mode.…”

Section: Biological Testsmentioning

confidence: 99%

Bhc‐cNMPs as either Water‐Soluble or Membrane‐Permeant Photoreleasable Cyclic Nucleotides for both One‐ and Two‐Photon Excitation

et al. 2004

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Cyclic nucleoside monophosphates (cNMPs) play key roles in many cellular regulatory processes, such as growth, differentiation, motility, and gene expression. Caged derivatives that can be activated by irradiation could be powerful tools for studying such diverse functions of intracellular second messengers, since the spatiotemporal dynamics of these molecules can be controlled by irradiation with appropriately focused light. Here we report the synthesis, photochemistry, and biological testing of 6-bromo-7-hydroxycoumarin-4-ylmethyl esters of cNMP (Bhc-cNMP) and their acetyl derivatives (Bhc-cNMP/Ac) as new caged second messengers. Irradiation of Bhc-cNMPs quantitatively produced the parent cNMPs with one-photon uncaging efficiencies (Phiepsilon) of up to one order of magnitude better than those of 2-nitrophenethyl (NPE) cNMPs. In addition, two-photon induced photochemical release of cNMP from Bhc-cNMPs (7 and 8) can be observed with the two-photon uncaging action cross-sections (delta(u)) of up to 2.28 GM (1 GM=10(-50) cm(4) s photon(-1)), which is the largest value among those of the reported Bhc-caged compounds. The wavelength dependence of the delta(u) values of 7 revealed that the peak wavelength was twice that of the one-photon absorption maximum. Bhc-cNMPs showed practically useful water solubility (nearly 500 microM), whereas 7-acetylated derivatives (Bhc-cNMPs/Ac) were expected to have a certain membrane permeability. Their advantages were demonstrated in two types of biological systems: the opening of cAMP-mediated transduction channels in newt olfactory receptor cells and cAMP-mediated motility responses in epidermal melanophores in scales from medaka fish. Both examples showed that Bhc and Bhc/Ac caged compounds have great potential for use in many cell biological applications.

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“…However, addition of uncharged aromatic chromophores decreases the solubility of the caged cyclic nucleotides (Table 1). To mitigate this problem a dicarboxylate moiety has been added to both the nitrobenzyl and DEAC caged-cAMP probes 21,22 . We adopted the same strategy by conjugation of additional charge to the core DEAC450 chromophore (Supplemental video and Scheme 1).…”

Section: Resultsmentioning

confidence: 99%

“…Thus, the solubility of DEAC450-caged cAMP was such that a solution of a concentration of at least 1 mM could be made in physiological buffer, without the addition of any organic co-solvent (see Supporting Information). In general, when the caged compound uses an ester linkage to attach the chromophore to the biomolecule the stability of the linkage is defined by its electrophilic character 21 . For example, dimethoxy- o -nitrobenzyl-cAMP is much more unstable than its simple nitrobenzyl counter part 20 , and an electron deficient analog is more stable 21 .…”

Section: Resultsmentioning

confidence: 99%

Spectral Evolution of a Photochemical Protecting Group for Orthogonal Two-Color Uncaging with Visible Light

et al. 2013

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Caged compounds are molecules rendered functionally inert by derivatization with a photochemical protecting group. We describe the design logic behind the development of a diethylaminocoumarin (DEAC) caging chromophore, DEAC450, that absorbs blue light strongly (ε450 = 43,000 M-1 cm-1), and violet light 10-fold more weakly. The absorption minimum is in the wavelength range (340-360 nm) that is traditionally used for photolysis of many widely used nitroaromatic caged compounds (e.g. 4-carboxymethoxy-5,7-dinitroindolinyl(CDNI)-GABA). We used this chromophore to synthesize DEAC450-caged cAMP and found this probe was very stable towards aqueous hydrolysis in the electronic ground state but was photolyzed with a quantum efficiency of 0.76. When DEAC450-cAMP and CDNI-GABA where co-applied to striatal cholinergic interneurons, the caged compounds were photolyzed in an chromatically orthogonal manner using blue and violet light so as to modulate the neuronal firing rate in a bi-directional way.

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Photolabile Precursors of Cyclic Nucleotides with High Aqueous Solubility and Stability

Cited by 23 publications

References 28 publications

Caged compounds: photorelease technology for control of cellular chemistry and physiology

Caged compounds: photorelease technology for control of cellular chemistry and physiology

Bhc‐cNMPs as either Water‐Soluble or Membrane‐Permeant Photoreleasable Cyclic Nucleotides for both One‐ and Two‐Photon Excitation

Spectral Evolution of a Photochemical Protecting Group for Orthogonal Two-Color Uncaging with Visible Light

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