Abstract
Photochemical uncaging of bio-active molecules was introduced in 1977, but since then, there has been no substantial improvement in the properties of generic caging chromophores. We have developed a new chromophore, nitrodibenzofuran (NDBF) for ultra-efficient uncaging of second messengers inside cells. Photolysis of a NDBF derivative of EGTA (caged calcium) is about 16–160 times more efficient than photolysis of the most widely used caged compounds (the quantum yield of photolysis is 0.7 and the extinction coefficient is 18,400 M−1 cm−1). Ultraviolet (UV)-laser photolysis of NDBF-EGTA:Ca2+ rapidly released Ca2+ (rate of 20,000 s−1) and initiated contraction of skinned guinea pig cardiac muscle. NDBF-EGTA has a two-photon cross-section of ∼0.6 GM and two-photon photolysis induced localized Ca2+-induced Ca2+ release from the sarcoplasmic recticulum of intact cardiac myocytes. Thus, the NDBF chromophore has great promise as a generic and photochemically efficient protecting group for both one- and two-photon uncaging in living cells.
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Acknowledgements
This work was supported by grants from the National Institutes of Health (GM53395), American Heart Association, McKnight Endowment Fund for Neuroscience, Human Frontiers Science Program, the Swiss National Science Foundation (3100-061344) and the PA Tobacco Fund.
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G.C.R.E.-D. and A.M. have filed an application for a US patent for NBDF.
Supplementary information
Supplementary Fig. 1
NDBF-EGTA Ca2+ titration. (PDF 53 kb)
Supplementary Fig. 2
NDBF-EGTA Mg2+ titration. (PDF 287 kb)
Supplementary Fig. 3
NDBF-EGTA aci-nitro intermediate decay. (PDF 55 kb)
Supplementary Fig. 4
Generic nature of NDBF photochemistry. (PDF 24 kb)
Supplementary Table 1
Summary of the photochemical properties of coumarin caged compounds. (PDF 27 kb)
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Momotake, A., Lindegger, N., Niggli, E. et al. The nitrodibenzofuran chromophore: a new caging group for ultra-efficient photolysis in living cells. Nat Methods 3, 35–40 (2006). https://doi.org/10.1038/nmeth821
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DOI: https://doi.org/10.1038/nmeth821
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