Journal of the American Chemical Society
COMMUNICATION
as recently suggested in the case of the zinc-responsive fluorescent
probes FluoZin-3 and TSQ.25 Despite its net anionic charge at
neutral pH, CTAP-2 proved to be cell permeant and produced in
live NIH 3T3 cells a perinuclear staining pattern (Figure S7),
reminiscent of the subcellular copper distribution previously
reported;5,26 however, in view of the above findings, the interpreta-
tion of the observed cellular staining is nontrivial and will require
further detailed studies.
Applicatons; Wang, B., Anslyn, E. V., Eds.; John Wiley & Sons: New
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In conclusion, we have developed a fluorescent probe CTAP-2
that selectively responds to Cu(I) in aqueous buffer with a
65-fold fluorescence enhancement. The response of CTAP-2 is
rapid and reversible, making it suitable as an indicator for titrations
with Cu(I) or for monitoringequilibrium concentrations of Cu(I).
While previously described fluorescent probes for Cu(I) have not
been reported to dissolve directly in water,4,5,9,10,27 salts of CTAP-
2 quickly dissolve in pure water up to millimolar concentrations,
circumventing the possibility of colloidal aggregate formation that
exists when organic stock solutions of poorly soluble dyes are
diluted into aqueous buffer. Because the majority of fluorescent
probes utilizedinbiology areconsiderablylipophilic, theformation
of colloids is likely not limited to the probes investigated here but a
rather widespread phenomenon that deserves particular attention
when interpreting fluorescence microscopy data. Adding to the
previously described applications of Cu(I)-responsive fluorescent
probes, CTAP-2 was able to detect copper bound to a metallo-
chaperone. Asonly proteinswithaccessible metal sites can give rise
to a fluorescence response, the detection of metalloproteins with
fluorescent indicators such as CTAP-2 nicely complements the
currently available techniques for in-gel metal profiling, namely
laser ablationÀinductively coupled plasma mass spectrometry
(LA-ICP-MS)28 and synchrotron-based X-ray fluorescence
mapping,29 both of which measure the total metal content
regardless of its accessibility, thus further expanding the metallo-
proteomics toolbox.30
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’ AUTHOR INFORMATION
(22) Robinson, N. J.; Winge, D. R. Annu. Rev. Biochem. 2010, 79, 537.
(23) Narindrasorasak, S.; Zhang, X.; Roberts, E. A.; Sarkar, B.
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Corresponding Author
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’ ACKNOWLEDGMENT
Financial support from the National Institutes of Health
(R01GM067169) is gratefully acknowledged. We thank Dr.
David L. Huffman for the gift of the hAtox1 expression vector,
and Jonathan Hofmekler for the preparation of CS1 and CS3.
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