POTENT INHIBITION OF RIBULOSE-BISPHOSPHATE CARBOXYLASE BY AN OXIDIZEDIMPURITY IN RIBULOSE-1,5-BISPHOSPHATE

Oxidation of D-ribulose-1,5-bisphosphate (ribulose-P-2) during synthes is and/or storage produces D-glycero-2,3-pentodiulose-1,5-bisphosphate (pentodiulose-P-2), a potent slow, tight-binding inhibitor of spinach (Spinacia oleracea L.) ribulose-P-2 carboxylase/ oxygenase (Rubisco). Differing degrees of contamination with pentodiulose-P-2 caused the d ecline in Rubisco activity seen during Rubisco assay time courses to v ary between different preparations of ribulose-P-2. With some ribulose -P-2 preparations, this compound can be the dominant cause of the decl ine, far exceeding the significance of the catalytic by-product, D-xyl ulose-1,5-bisphosphate. Unlike xylulose-1,5-bisphosphate, pentodiulose -P-2 did not appear to be a significant by-product of catalysis by wil d-type Rubisco at saturating CO2 concentration, it was produced slowly during frozen storage of ribulose-P-2, even at low pH, more rapidly i n Rubisco assay buffers at room temperature, and particularly rapidly on deliberate oxidation of ribulose-P-2 with CU2+. Its formation was p revented by the exclusion of transition metals and O-2. Pentodiulose-P -2 was unstable and decayed to a variety of other less-inhibitory comp ounds, particularly in the presence of some buffers. However, it forme d a tight, stable complex with carbamylated spinach Rubisco, which cou ld be isolated by gel filtration, presumably because its structure mim ics that of the enediol intermediate of Rubisco catalysis. Rubisco cat alyzes the cleavage of pentodiulose-P-2 by H2O2, producing P-glycolate .