The crystal structure of the Escherichia coli lactose permease at 3.5
Å with a bound substrate has been reported recently. The structure
reveals the sugar–protein contacts, which include hydrophobic
stacking between the galactopyranosyl ring of substrate and the
indole side chain of Trp-151, as proposed previously. The nature of
this interaction is studied here by exploiting the luminescence
properties of Trp-151 in a mutant devoid of other tryptophan
residues. The following phenomena are observed. (i) The fluorescence
emission spectrum of Trp-151 and fluorescence-quenching
experiments with water-soluble quenchers demonstrate that Trp-
151 is in a hydrophilic environment. (ii) Substrate binding leads to
a blue shift in the emission spectrum and reduction in accessibility
to polar quenchers, indicating that Trp-151 becomes less exposed
to aqueous solvent. (iii) The phosphorescence spectrum of Trp-151
is red-shifted in the presence of substrate, indicating charge
separation of the triplet state due to a direct stacking interaction
between the galactopyranosyl and indole rings. The spectroscopic
data fully complement the x-ray structure and demonstrate the
feasibility of fluorescence spectroscopy for studying sugar–protein
interactions.