It should be noted that the electric field around the fluorophore in the protein cavity is not uniform and quite different from the local electric field in a polar solvent. An approach to take into account the nonuniformity of the electrostatic environment is applying an oriented external electric field. The insight obtained above with implicit solvent model shows how a homogeneous dielectric environment affects the fluorescence spectrum. However, investigating the effect of OEEFs is more useful for capturing the heterogeneous electrostatic effects and predicting how the change at a particular position (such as the mutation of a specific amino acid of the protein) may influence the spectrum. The OEEFs were applied along three mutually perpendicular directions, as illustrated in Scheme 2. The fluorescence wavelength (λ F ) was found to be most sensitive to the OEEFs along the X axis (F X ), but much less sensitive to the fields along the Y and Z axes (refer to Figures and for the effect of Y and F Z , respectively). Applying a positive F X field leads to a blueshift in fluorescence wavelength, while a negative F X gives rise to a redshift (Figure ). The spectral shifting directly correlates with the field strength, stronger fields resulting in larger offsets. Specifically, a F X field of -0.0075 a.u. reduces the vertical emission energy (E F ) by 0.63 eV from 3.01 eV to 2.38 eV, shifting λ F from 412 nm to 521 nm. Contrary to above case of polar solvents, the fluorescence intensity was found to be insensitive to F X (Figure ). For F Y , positive fields result in blueshifts, while negative fields result in redshifts. The effect of F Y is much smaller than that of F X . Applying a negative field of F Y = -0.0075 a.u. reduces E F by only 0.20 eV from 3.01 eV to 2.81 eV, red-shifting λ F by 29 nm from 412 nm to 441 nm. In contrast, positive F Z fields redshift the wavelength while negative F Z fields lead to blueshifts. The decrease of E F induced by F Z = 0.0075 a.u. is only 0.13 eV (from 3.01 eV to 2.88 eV), corresponding to an offset of only 9 nm for λ F . Since there is a small difference between the TD-DFT/MMoptimized geometry in this study and the geometry previously obtained by some of us using the multi-reference method, we also tested the effect of OEEFs using this geometry. The results are collectively shown in Figures . Overall, the effects of OEEFs are totally consistent with the results discussed above, further validating the robustness of our results (refer to Note 1 in the SI).