Duetta

The samples were dissolved or dispersed in ultra-pure water to 0.02 wt% and measured using a fluorospectrometer (Duetta, Horiba, Kyoto, Japan). The supplied software (EzSpec) was used for analysis. Sample solutions were placed into quartz cuvettes with an optical path length of 1 cm. Measurements were taken with stirring at 1000 rpm. The excitation wavelength was set at 280 nm for tryptophan quenching. Emission spectra were recorded in the range between 300 and 400 nm. Excitation and emission slits were set at 5 nm. All spectra were the average of at least three repeated scans.

The morphologies of the HepG2 cells were observed by SEM (Hitachi, Japan). The morphologies of the Au nanoparticles were measured by TEM (JEM-2100F, JEOL). The FL, FT-IR, XPS and UV-vis absorption spectra of Au nanoparticles were obtained by a fluorescence spectrometer (Duetta, HORIBA), Fourier transform infrared spectrometer (Nicolet iS5, Thermo Fisher Scientific), x-ray photoelectron spectrometer (ESCALAB 250, Thermo Fisher Scientific), and UV-vis spectrophotometer (UV6100, MAPADA). The dark-field image and fluorescence image of HepG2 cell were measured by CLSM (A1R HD25, Nikon). The images of pathological sections were captured using a fluorescence microscope (Axio Imager 2, ZEISS). The element content of HepG2 cell was investigated with inductively coupled plasma mass spectrometer (Ultima Expert LT, HORIBA). The microplate reader (Multiskan, Thermo Fisher Scientific) was used to measure cell viability. Gel imaging system is used for Western blot image (E-Gel Imager, Thermo Fisher Scientific). The NMR spectra of auranofin before and after treatment were tested by an NMR spectrometer (Avance 500 MHz, Bruker).

For labelling, the purified NUP98 FG domain with single or double cysteine mutations was exchanged to 4 M GdmCl, 1× PBS, 0.1 mM EDTA and 0.2 mM TCEP, pH 7. Labelling with Alexa Fluor 594 maleimide (A10256, Thermo Fisher) and LD655-maleimide (Lumidyne Technologies) was done at the molar ratio of 1:2 (dye:protein) overnight at 4 °C. The reaction was quenched with 10 mM DTT in 4 M GdmCl and 1× PBS, pH 7. Unreacted dye was washed off using a 3-kDa MWCO centrifugal filter, and the labelled protein was further purified with Superdex 200. Pure fractions were chosen, pooled and concentrated as described above, and the final concentration was measured by the absorbance spectrometer Duetta (Horiba). The proteins were flash-frozen and stored at −80 °C.