SULFITOLYSIS AND THIOREDOXIN-DEPENDENT REDUCTION REVEAL THE PRESENCE OF A STRUCTURAL DISULFIDE BRIDGE IN SPINACH CHLOROPLAST FRUCTOSE-1,6-BISPHOSPHATASE

A significant difference between cytosolic and chloroplastic fructose- 1,6-bisphosphatase (FbPase) is an extra peptide in the middle of chlor oplast FbPase which contains three additional cysteine residues. Sit-d irected mutagenesis experiments have shown that at least two of these cysteine residues are involved in forming the regulatory disulfide bri dge [Jacquot, J.-P. et al., FEBS Lett. 401 (1997) 143-147] which is th e presupposition for the thioredoxin-dependent control of chloroplast FbPase activity. Here we report that each subunit of the FbPase contai ns an additional structural disulfide bridge which has been observed b y combined application of thioredoxins and sulfitolysis. Observation o f the structural disulfide bridges by sulfitolysis was only possible w hen the FbPase was already specifically reduced by the homologous thio redoxin species TR, and TRf from spinach chloroplasts. Interestingly, the accessibility of the structural disulfide bridge for sulfite ions depends on the thioredoxin species engaged in the thioredoxin/FbPase c omplex. (C) 1998 Federation of European Biochemical Societies.