Fig. 4: d-Cys does not support sulfur transfer within NFS1 during de novo [2Fe–2S] cluster biosynthesis.
From: d-cysteine impairs tumour growth by inhibiting cysteine desulfurase NFS1
a, Enzymatic [2Fe–2S] cluster reconstitution by the core ISC complex on the ISCU2 scaffold using l-Cys (positive control), d-Cys and mixtures of both enantiomers as indicated. Final Cys concentration was 1 mM for all mixtures, except for 10× and 20× d-Cys (0.5 mM). b,c, Cys-ketimine generation on (NIA)2, (NIAX)2 and (NIAUX)2 complexes measured by UV/Vis spectroscopy. The increase in the absorption at 340 nm indicates the generation of the l-Cys-ketimine (b) and d-Cys-ketimine (c) at the expense of the internal aldimine absorbing around 416 nm. Nearly identical rates of Cys-ketimine formation were observed for l-Cys and d-Cys (for time courses see Extended Data Fig. 7b,c). (NIA)2 spectra were recorded every 4 min, while (NIAX)2 and (NIAUX)2 spectra were recorded every 90 s. d, Persulfidation of Cys381NFS1 (NFS1 + 6) in the presence of l-Cys or d-Cys or mixtures thereof at the indicated concentrations (for explanation of the assay see Supplementary Fig. 7; representative image of two biological replicates is shown). d-Cys did not enable any NFS1 persulfidation (NFS1 + 7). The left lane shows non-labelled NFS1 as a control. The band at 70 kDa is the Escherichia coli DnaK chaperone, which does not contain a persulfide. a.u., arbitrary units.
