Further studies on the quaternary structure of D-ribulose-1, 5-bisphosphate carboxylase from Alcaligenes eutrophus.
Homogeneous D-ribulose-1,5-bisphosphate carboxylase, isolated from the hydrogen bacterium Alcaligenes eutrophus, has been studied by analytical ultracentrifugation. Sedimentation equilibrium experiments showed the enzyme to have a molecular weight M c =0 r =534000. The sedimentation coefficient was S0(20); w = 14.1S. The two types of subunits constituting the ribulosebisphosphate carboxylase were separated by gel filtration in the presence of sodium dodecylsulphate and the amino acid compositions of the isolated large and small subunits were determined. Rabbit antibodies were developed against the ribulosebisphosphate carboxylase and its isolated subunits. The specific reactivity of the respective antibodies with their homologous antigens was proven by double immunodiffusion and quantitative immunoprecipitation analyses. Antibodies elicited against the whole enzyme also reacted with both the isolated large and small subunits as did the subunit-specific antibodies with the whole enzyme. There was no immunological correspondence between the large and the small subunits. The specific inhibition of the enzyme activity by antibodies directed against sites on the large subunit suggests that the catalytic function resides in the large subunit. Electron microscopic examination of antibody . carboxylase complexes formed upon mixing of the specific immunoglobulins G with the enzyme was used to verify the arrangement of the large and small subunits in our recently proposed structural model of the enzyme molecule. The results confirmed that the large subunits are located in the central two layers of the four-layered enzyme molecule, whereas the two outer layers consist of small subunits. The observations are discussed with respect to an alternative model for the quaternary structure of ribulosebisphosphate carboxylase from tobacco.
D000421:Alcaligenes / Q000201:enzymology*; D000596:Amino Acids / Q000032:analysis; D000936:Antigen-Antibody Complex; D002262:Carboxy-Lyases; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008970:Molecular Weight; D011487:Protein Conformation; D012273:Ribulose-Bisphosphate Carboxylase / Q000276:immunology
D000596:Amino Acids; D000936:Antigen-Antibody Complex; D046911:Macromolecular Substances; D002262:Carboxy-Lyases; D012273:Ribulose-Bisphosphate Carboxylase