Partition kinetics of ribulose-1,5-bisphosphate carboxylase from Rhodospirillum rubrum.
When the enzymatically generated intermediate 2-carboxy-3-keto-D-arabinitol-1,5-bisphosphate (II) was used as a substrate with fresh enzyme, 70% reacted to produce 3-phosphoglycerate (3PGA). When a reaction mixture of enzyme plus [1-32P]ribulose 1,5-bisphosphate (RuBP) was quenched in the steady state with the tightly bound inhibitor 2-carboxyarabinitol-1,5-bisphosphate, 30% of the enzyme-bound species was released as 3PGA and 70% as RuBP. The major source for this partition was the ternary substrates Michaelis complex. The level of carboxylated intermediate in the steady state was determined to be 8% of active sites under the conditions of substrate saturation. No burst was seen in the appearance of product when 6.5 eq of [1-32P]RuBP was mixed with enzyme plus saturating CO2 and the reaction followed in the steady state. From these data plus the steady-state Vmax and Km of RuBP it is possible to derive the five bulk rate constants represented in the scheme ECO2 + RuBP in equilibrium ERuBPCO2 in equilibrium E X II----E + 2(3PGA).
D005988:Glyceric Acids / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives / Q000378:metabolism; D007700:Kinetics; D008433:Mathematics; D012247:Rhodospirillum rubrum / Q000201:enzymology*; D012273:Ribulose-Bisphosphate Carboxylase / Q000378:metabolism*
D005988:Glyceric Acids; C027494:N,N-bis(2-hydroxyethyl)glycine; C005156:3-phosphoglycerate; D012273:Ribulose-Bisphosphate Carboxylase; D005998:Glycine