title: Three partial reactions of ribulose-bisphosphate carboxylase require both large and small subunits.
abstract: Three partial reactions of ribulose-bisphosphate carboxylase/oxygenase were measured in the presence and absence of small subunits using the enzyme from the cyanobacterium, Synechococcus ACMM 323, whose small subunits may be reversibly dissociated from its octameric, large-subunit core. These partial reactions were: the exchange of the proton at C-3 of the substrate, ribulose 1,5-bisphosphate, with the medium which is indicative of C-2, C-3 enolization; the hydrolysis of the 6-carbon reaction intermediate, 3-keto-2-carboxy-D-arabinitol 1,5-bisphosphate, to two molecules of 3-phosphoglycerate; and the decarboxylation of the 6-carbon intermediate, which is catalyzed only by the deactivated, divalent metal-ion-free carboxylase. None of these partial reactions was catalyzed by the small-subunit-depleted, large-subunit octamer to an extent greater than that expected from the residual small subunit content (about 3%), implying that small subunits are required for all three reactions. Clearly, the small subunit's influence is not restricted to any single stage of the catalytic sequence. Under conditions where it was possible to demonstrate tight binding of the reaction-intermediate analog, 2-carboxy-D-arabinitol 1,5-bisphosphate, to the large-subunit octamer, no binding of the 6-carbon intermediate could be detected. We suggest that either the tight-binding form of the 6-carbon intermediate is the hydrated gem-diol, not the ketone, or the large subunits by themselves intrinsically possess a trace of catalytic activity which discharges any bound intermediate before it can be measured.
mesh_terms: D002245:Carbon Dioxide / Q000378:metabolism; D002850:Chromatography, Gel; D000458:Cyanobacteria / Q000201:enzymology; D007700:Kinetics; D046911:Macromolecular Substances; D008274:Magnesium / Q000378:metabolism; D008433:Mathematics; D008956:Models, Chemical; D011522:Protons; D012273:Ribulose-Bisphosphate Carboxylase / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D046911:Macromolecular Substances; D011522:Protons; D002245:Carbon Dioxide; D012273:Ribulose-Bisphosphate Carboxylase; D008274:Magnesium