Journal
CHEMBIOCHEM
Volume 7, Issue 4, Pages 669-672Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.200500515
Keywords
apoptosis; excimers; fluorescent probes; pyrene; TUNEL
Funding
- NIGMS NIH HHS [GM067201] Funding Source: Medline
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A number of research laboratories have investigated the properties of multichromophore molecules and their applications in materials science and in biotechnology. Previous approaches for preparing such molecules have involved traditional organic synthesis. Here we describe the one-step enzymatic synthesis of such a multichromophore species by using a DNA-polymerizing enzyme (terminal deoxynucleotidyl transferase (TdT)). We find that a nucleotide-like molecule with pyrene replacing the DNA base (dPTP) can be accepted as a substrate for this enzyme to produce discrete chromophores that have 3 or 4 pyrenes consecutively, depending on which anomer (alpha or beta) is used. Products were characterized by gel electrophoresis, mass spectrometry, and fluorescence. The reaction was found to change the fluorescence emission of the chromophore from a maximum at 375 nm (the monomer nucleotide) to 490 nm in the oligomeric product. This new green-white emission is consistent with the formotion of a pyrene excimer between adjacent pyrene glycosides, which exhibit a large Stokes shift of 130 nm. The enzymatic synthesis of the pyrene excimer might hove applications in homogeneous biological assays for DNA fragments, such as those that arise during apoptosis.
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