FORMATION OF THE ACTIVE-SITE OF RIBULOSE-1,5-BISPHOSPHATE CARBOXYLASEOXYGENASE BY A DISORDER ORDER TRANSITION FROM THE UNACTIVATED TO THE ACTIVATED FORM

Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) catalyzes th e key first step in photosynthetic CO2 fixation, the reaction that inc orporates CO2 into sugar. In this study, refined crystal structures of unactivated tobacco RuBisCO and activated RuBisCO from spinach and to bacco, in complex with the reaction-intermediate analog 2-carboxyarabi nitol 1,5-bisphosphate (CABP), are compared. Both plant enzymes are he xadecameric complexes of eight large and eight small subunits with a t otal relative molecular mass of almost-equal-to 550,000. The compariso n of activated and unactivated forms of RuBisCO provides insight into the dynamics of action of this enzyme. The catalytic site, which is op en to the solvent in the unactivated enzyme, becomes shielded in the a ctivated CABP complex. This shielding is accomplished by a 12-angstrom movement of the active-site ''loop 6'' (residues 331-338) and a disor der-order transition of three loops near the active-site entrance, the N terminus, the C terminus, and a loop comprising residues 64-68. All these residues belong to the catalytic large subunit. Domain rotation s of about 2-degrees are observed, also tightening the active-site cle ft. These observations provide an explanation for the extremely tight binding (K(d) less-than-or-equal-to 10(-11)M) of the CABP molecule. A striking correlation exists between crystallographic temperature facto rs in the activated enzyme and the magnitude of the atomic movement up on activation.