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. Author manuscript; available in PMC: 2013 Jan 21.
Published in final edited form as: Methods Mol Biol. 2012;796:133–174. doi: 10.1007/978-1-61779-334-9_8

Figure 11. Intrinsic Tryptophan Fluorescence of Heterologously Expressed G3 subunits.

Figure 11

The spectrum shows the fluorescence signal from ~200 nM of purified wild-type Gαi1 in a buffer containing 50 mM Tris-Cl pH 8.0, 50 mM NaCl, 2 mM MgCl2, 1 mM DTT, and 10 μM GDP before and after the addition of AIF4−. Purified protein added to Storage Buffer (50 mM Tris-Cl pH 8.0, 50 mM NaCl, 2 mM MgCl2, 1 mM DTT, and 10 μM GDP) results in basal fluorescence arrow at 1.3 minutes). AlF4− is added by bringing the concentration of NaF in the cuvette to 10 mM and AlCl3 to 50 μM. Binding of to GDP-Gα results in an increase in the fluorescence from Trp211 (arrow at 2 minutes). The increase in fluorescence is calculated as the percentage difference in the average fluorescence signal of GDP- versus GDP-AlF4−-bound Gαi1.