Figure 1.
Relative orientations of the side-chains at position 130 and its long-range interaction partners in β-barrel-α-helix interface of TL.
A. Relative orientation of native Phe130 (χ1 = 180°). B. The rotamers t (χ1 = 180°) and g− (χ1 = −60°) of Trp at position 130. The mutation to introduce Trp at position 130 and rotamer assignment were generated from the PDB file (1XKI) with DS Visualizer 3.0 (Accelrys Inc.).
Figure 2.
Far-UV CD spectra of the mutants with single Trp130 in which long-range interaction (the positions 113 and 115) were modified.
The CD spectrum of native TL is shown for comparison. For clarity the spectra are divided in two sets (A) and (B).
Table 1.
Secondary structure content for TL mutants at pH value of 7.3 determined from CD spectra.
Figure 3.
The corrected fluorescence spectra of the mutants with single Trp130 in which long-range interaction (the positions 113 and 115) were modified.
Fluorescence intensities were normalized to the same absorbance (0.05) at 295 nm to reflect respective quantum yield values. For clarity the spectra are divided in two sets (A) and (B).
Figure 4.
Representative phase angle (open symbols) and modulation (solid symbols) fluorescence lifetime data of TL mutant W130, W130F113F115 and W130A113 at pH 7.3.
Solid lines represent the best bi- or tri-exponential fit for the parameters given in Table 1.
Table 2.
Fluorescence lifetime parameters for the mutants with single Trp130 located in the α-helix of human tear lipocalin in various long-range interacting partners located in the positions 113 and 115.