Extended Data Fig. 7. AoMoeZ-dependent protein thiocarboxylate formation of sulphur carrier proteins using thiosulphate as the sulphur source.
a–d, Deconvoluted ESI-MS of (a) AoMoaD2 incubated with AoMoeZ (the observed peaks are consistent with the calculated molecular masses of N-His6-AoMoaD2-COSH (12489), N-His6-AoMoaD2-glycerol (12547), and N-gluconoylated-His6-AoMoaD2-COSH (12667)), (b) AoMoaD2 incubated with AoMoeZ(C360A) mutant (the observed peaks are consistent with the calculated molecular masses of N-His6-AoMoaD2 (12473) and N-His6-AoMoaD2-glycerol (12547)), (c) AoCysO incubated with AoMoeZ (the observed peaks are consistent with the calculated molecular masses of N-His6-AoCysO-COSH (11704), N-His6-AoCysO-glycerol (11762), and N-gluconoylated-His6-AoMoaD2-COSH (11882)), and (d) AoCysO incubated with AoMoeZ(C360A) mutant (the observed peaks are consistent with the calculated molecular masses of N-His6-AoCysO (11688), N-His6-AoCysO-glycerol (11762), their N-gluconoylated derivatives (11866, and 11940), N-His6-AoCysO-AMP (12017)). Observed masses corresponding to protein thiocarboxylate are shown in red. Two peaks corresponding to the dehydration of N-His6-AoMoaD2 and N-His6-AoCysO were likely caused by in-source CID during the ESI-MS analysis. e, Kinetic parameters for the thiosulphate: cyanide sulphurtransferase activity of MoeZ from A. orientalis. Bovine liver rhodanese is a typical rhodanese enzyme. Compared to the bovine rhodanese, human molybdopterin synthase sulphurase (human MOCS3) displayed much lower thiosulphate:cyanide sulphurtransferase activity. In the case of human MOCS3, L-cysteine and cysteine desulphurase are proposed as the physiological sulphur source over thiosulphate because of its lower rhodanese activity43,44. However, this may not be the case for MoeZ from A. orientalis (AoMoeZ) since its rhodanese activity is comparable to bovine liver rhodanese as shown above.