Figure 1: Principle of hole-burning
spectroscopy
A Schematic description
of inhomogeneous broadening. The top panel contains depicts the
absorption spectrum of a single molecule, with a characteristic
(homogeneous) absorption linewidth Ghom. The
next panel depicts the spectra of 10 molecules; each showing slightly
different absorption line centers due to differences in their
interactions with the host matrix. The next two panels depicts
the case for 103 and 106 molecules respectively.
In the latter case, the total absorption spectrum is a sum of
the individual spectra for each molecule and is characteristically
(inhomogeneously) broadened with linewidth -Ginhom.
B Schematic description of an inhomogeneously
broadened absorption line (center wavelength l0) and
the hole burned at the wavelength of laser excitation (l1).
The photoproduct (absorbing at a different wavelength) is also
depicted. C (Left Panel): Excitation spectrum
of wild-type GFP at 1.6 K before (æ) and after (---) burning
a hole into A at 434±1 nm. The insets in the top left and
bottom right of the panel are typical holes burned into the A
and I states respectively. The hole widths ~ 2 cm -1
are limited by the laser bandwidth. The fundamental absorption
transitions are determined to be A0-0 = 434±1
nm; I0-0 = 495±1 nm. (Right Panel): Corresponding
emission spectrum of I upon direct excitation of I at 493 nm.
The I state is produced upon burning into A, clearly demonstrating
the photoconversion A-> I. The GFP was dissolved in a buffer/glycerol
solution that transforms into a glass at cryogenic temperatures.
Data taken from (3).
