Fig. 2.
Effect of sugar binding on fluorescence of DACM- or MIANS-labeled LacY. (A) Overall structure of LacY [Protein Data Bank (PDB) ID code 1pv7] with TDG occupying the sugar-binding site in the middle of the molecule at the apex of the hydrophilic cavity opened to the cytoplasmic side. The black sphere represents Cα atom of position 331 where the V331C mutation was introduced. LacY structure is displayed by using Pymol 0.97 (DeLano Scientific). (B and C) Structures of fluorophores and emission spectra of V331C LacY labeled with DACM or MIANS, respectively. Solid line, no sugar added or 10 mM sucrose; broken line, 10 mM TDG added. The spectra were recorded at 0.4 μM protein in 50 mM NaPi (pH 7.5), 0.02% DDM with excitation wavelengths of 397 nm or 330 nm for DACM- or MIANS-labeled LacY, respectively.