The Fluorescence Resonant Energy Transfer (FRET) Gate: A Time-Resolved Study

Alan J. Heeger (UC Santa Barbara), DMR-0099843

The two-step energy transfer process in a self-assembled complex comprising a cationic conjugated polymer (CCP) and a double stranded DNA (dsDNA) was investigated using pump-dump-emission spectroscopy and time correlated single photon counting; energy is transferred from the CCP to an ethidium bromide (EB) molecule intercalated into the dsDNA via a fluorescein molecule linked to one terminus of the DNA. Time-dependent anisotropy measurements indicate that the inefficient direct energy transfer from the CCP to the intercalated EB results from the near orthogonality of their transition moments. These measurements also show that the transition moment of the fluorescein spans a range of angular distributions that lies between that of the CCP and EB. Consequently, the fluorescein acts as a FRET gate to relay the excitation energy from the CCP to the EB.

Schematic representation of two-step FRET from CCP to dyes intercalated into DNA in the absence (A) and presence (B) of the FRET gate. The molecular structures of the cationic conjugated polymer (PFP), fluorescein (FL) and ethidium bromide (EB) are also shown.

Relative orientations of the three optical components:conjugated polymer (PFP, donor), fluorescein,FL)and intercalated EB (acceptor).

The Fluorescence Resonant Energy Transfer (FRET) Gate: A Time-Resolved Study

Alan J. Heeger (UC Santa Barbara), DMR-0099843

Education

This is a classic interdisciplinary study involving sophisticated ultrafast time resolved pulsed laser spectroscopy on a self-assembled structure of direct interest and importance to biology. The work was carried out largely by post-doctoral researchers with previous research backgrounds narrowly focused in chemistry. The interdisciplinary nature of the research was a stimulating educational experience. The demonstration of the details of the time-resolved anisotropic energy transfer demonstrates the reality of the schematic diagrams shown on the first slide. Although Förster energy transfer has been known for >50 years, this is the first example of time-resolved anisotropic FRET.

Outreach• The use of FRET on this self-assembled

complex between the DNA double helix is part of our ongoing studies directed toward detection of specific sequences on DNA. A company has been formed in Santa Barbara, Sirigen Inc., to pursue this approach to sequence detection on DNA (AJH is not involved in the company). This capability of direct detection of specific targeted sequences is of direct relevance to the prevention of bio-terrorism.

• Professor Heeger has given invited (Keynote and Plenary) lectures on the results of this research at conferences in the United States, Europe and Asia.