The transition between the open and closed states of rubisco is triggered by the inter-phosphate distance of the bound bisphosphate

D-Ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) catalyses the c entral CO2-fixing reaction of photosynthesis in a complex, multiple-step pr ocess. Several structures of rubisco complexed with substrate analogues, in hibitors and products have been determined by X-ray crystallography. The st ructures fall into two well-defined and distinct states. The active site is either "open" or "closed". The timing and mechanism of the transition betw een these two states have been uncertain. We solved the crystal structure o f unactivated (metal-free) rubisco from tobacco with only inorganic phospha te bound and conclude that phosphate binding per se does not trigger closur e, as it does in the similarly structured enzyme, triosephosphate isomerase . Comparison of all available rubisco structures suggests that, instead, th e distance between the terminal phosphates (P1 and P2) of the bisphosphate ligand is the trigger: if that distance is less than 9.1 Angstrom, then the active site closes; if it is greater than 9.4 Angstrom then the enzyme rem ains open. Shortening of the inter-phosphate distance results from the liga nd binding in a more curved conformation when O atoms of the ligand's sugar backbone interact either with the metal, if it is present, or with charged groups in the metal-binding site, if the metal is absent. This shortening brings the P1 phosphate into hydrogen bonding contact with Thr65. Thr65 exi sts in two discrete states related by a rotation of the backbone psi torsio n angle. This rotation is coupled to domain rotation and hence to active si te closure. Rotation of the side-chain of Thr65 also affects the C-terminal strand of large subunit which packs against Loop 6 after closure. The posi tion of the C-terminal strand in the closed state is stabilised by multiple polar interactions with a distinctive highly-charged latch site involving the side-chain of Asp473. Ln the open state, this latch site may be occupie d instead by phosphorylated anions. (C) 2000 Academic Press.