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Inhibition of ribulose bisphosphate carboxylase by substrate ribulose 1,5-bisphosphate.

1983; Elsevier BV; Volume: 258; Issue: 22 Linguagem: Inglês

10.1016/s0021-9258(17)43982-2

1083-351X

Douglas B. Jordan, Raymond Chollet,

Coenzyme Q10 studies and effects

Substrate ribulose bisphosphate is a potent and a weak inhibitor of the rate of CO2/Mg2+ activation in the carboxylase purified from spinach leaves and Rhodospirillum rubrum, respectively.At 2 "C, the concen- tration of ribulose bisphosphate required for 50% inhibition of the initial rate of C02/Mg2+ activation was less than 0.4 I.IM for the spinach enzyme, but between 67 and 270 I.IM for the R. rubrum carboxylase.Activator 14C02 trapping experiments demonstrated that ribulose bisphosphate inhibits activation by excluding activator COz from the spinach enzyme.The reason for the different sensitivities to inhibition by substrate was evident from equilibrium binding studies with the inactive enzyme forms which indicated that the K D (ribulose bisphosphate) was 0.02 1 p~ for spinach enzyme and 5.9 p~ for the R. rubrum protein.Inhibition of activation, however, was not explained by the equilibrium binding results alone.Ribulose bisphosphate was observed to dissociate very slowly from the inactive spinach enzyme (at 2 "c, koFF = 4.9 X s-').The release of substrate from the inactive R. rubrum carboxylase was much more rapid, with a minimum value for keto,, estimated at 5 X s-' at 2 "C.We conclude that strong inhibition of COZ/Mg2+ activation in the spinach enzyme is mediated by the tight binding and slow release of ribulose bisphosphate, which prevent activator COz and Mg2+ from binding to the protein.Weak inhibition of activation in the R. rubrum enzyme results from a larger K D value and a more rapid exchange of ribulose bisphosphate, which allow activator COz and Mg2+ to bind to the free enzyme between successive substrate-binding events.RuBP' carboxylase (EC 4.1.1.39)is present in all photosynthetic organisms, where it is a catalyst initiating both the photosynthetic carbon reduction and photorespiratory carbon oxidation cycles (1-4).The enzyme from all higher plants and most microorganisms is a hexadecamer comprised of 8 large subunits (Mr E 56,000) and 8 small subunits (Mr g 14,000).The catalytically essential amino acid residues (5-7) and the site of C02/Mg2+ activation (8) are assigned to the large subunit of the enzyme, and no clearly defined function of the