Development of a Single-Photon Spectrometer/Microscope: Probing Conformational Dynamics of Single Biomolecules on a Photon by Photon Basis

Harvard University, Cambridge MA

Investigators

Abstract

This award supports the construction of a new, state-of-the-art single-photon spectrometer/microscope. The instrument will enable study of the conformation of single molecules of enzymes and nucleic acids using photo-induced electron transfer techniques that have been pioneered by the PI. In comparison to available instruments, the device will have an extended excitation frequency range and significantly improved time resolution. The excitation makes use of a frequency-doubled and synchronously-pumped optical parametric oscillator. Through collaboration with an Italian group, a custom avalanche photodiode with a very small active area will be developed. The detector will permit the probing of short-distance conformational fluctuations. Among the types of experiments the instrument will permit are the use of photo-induced electron transfer between tetramethlyrodamine (TMR) dye and guanine to probe the conformational dynamics of double-stranded DNA, and the use of fluorescence resonant energy transfer (FRET) to probe dynamics of a DNA hairpin. The new instrument is expected to make a variety of other new single-molecule experiments possible, and to improve knowledge of macromolecular dynamics and of the role of conformational fluctuation on macromolecular function. The funds provided by the award will also support multidiciplinary training of graduate students and postdoctoral fellows working at the interface of physics, chemistry and biology.