Water-soluble through-bond energy transfer cassettes for intracellular imaging.
A special, water-soluble, fluorescent probe 1 was designed. This consisted of a fluorescein-based component to harvest irradiation at 488 nm and a rhodamine-based part designed to emit it at a significantly longer wavelength. This cassette was used to label an illustrative protein called ACBP. Evidence was accumulated to support the assertion that ACBP-1 bound its native ligand with a binding constant similar to that of the unlabeled protein, and retained its secondary structure (CD). ACBP-1 was imported into cells using the Chariot peptide. Confocal images proved that some ACBP-1 localized into the nucleus (as expected) and, most significantly, it could be visualized more effectively by irradiating at the donor (fluorescein-like) part of the cassette, than the acceptor (rhodamine-like) part. Overall, this study demonstrates that cassettes of this kind can label a protein without significantly perturbing its function or secondary structure and they can be visualized effectively via irradiation of the donor and observation of the acceptor fluorescence.
Bandichhor
Rakeshwar
R
Department of Chemistry, Texas A and M University, Box 30012, College Station, Texas 77841, USA.
Petrescu
Anca D
AD
Vespa
Aude
A
Kier
Ann B
AB
Schroeder
Friedhelm
F
Burgess
Kevin
K
D004735:Energy Transfer; D015394:Molecular Structure; D011506:Proteins / Q000737:chemistry*; D014867:Water / Q000737:chemistry*
D011506:Proteins; D014867:Water