Ribulose 1,5-diphosphate carboxylase (RUDPcase) from autotrophically grown Euglena gracilis was purified to homogeneity as measured by analytical ultracentrifugation, polyacrylamide gel electrophoresis, and immunoprecipitation reactions. The enzyme represented about 9% of total protein and 24% of soluble protein in the autotrophic cell. Light-grown, heterotrophic cells seemed to contain considerably less RUDPcase. Native carboxylase from autotrophic Euglena showed an s(20, w) at low protein concentrations of 17 to 17.5, suggesting a molecular weight of >500,000 daltons. Upon denaturation, the enzyme dissociated into two subunits having different amino acid compositions and molecular weights of 59,000 and 12,000 daltons. Based upon the amino acid mass ratios, a quaternary organization of 7 to 8 large and 8 to 10 small subunits per native enzyme molecule was indicated.The phylogenetic relationship of carboxylase from Euglena and from three higher plants was investigated. In general, the size, subunit formation, and quaternary structure of RUDPcase from the various sources seemed to be similar. A partial immunochemical reaction between anti-RUDPcase serum from Euglena and the enzymes from lettuce, cucumber, and New Zealand spinach suggested that the algal and higher plant carboxylases were related but not identical. This was borne out by amino acid analyses which showed a close correspondence between the large, but not the small, subunits of Euglena and lettuce.