For single molecule Förster resonance energy transfer (smFRET) experiments, total internal reflection fluorescence (TIRF) and confocal modalities were used to probe the FRET network consisting of 12 complementary pairs of FRET labeling sites. Comparing the two approaches provides validation through the use of different dyes, sample preparation, instrumentation, and handling. Confocal smFRET with pulsed interleaved excitation and multiparameter fluorescence detection were holistically analyzed in four modalities, including fluorescence decay analysis, two-dimensional representations, photon distribution analysis, and filtered fluorescence correlation spectroscopy. Each approach provides complementary information on the underlying conformational distribution with respect to inter-dye distances and protein dynamics. For simulations: replica-exchange discrete molecular dynamics (rxDMD) provided an independent view of the energy landscape. For structural modeling: two approaches were used. Rigid body docking used two fixed structures (PDZ3 and SH3-GuK) to identify domain orientations that satisfied the FRET-derived interdye distances for all 12 FRET pairs, resulting in a fully FRET-restrained model. Screening of the rxDMD simulations with the FRET distances and the boundaries from FRET robustness analysis allowed classification of structures that mutually satisfy the simulated model (rxDMD) and the FRET observables. Finally, to validate the proposed structural models, we used disulfide mapping and probed interfaces corresponding to the preferred structural basins. In a refinement step, we introduced the disulfide information for both structural modeling approaches.