Purification and species distribution of rubisco activase

Abstract

Ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) activase, a soluble chloroplast protein which promotes light-dependent rubisco activation, was partially purified from spinach chloroplasts by ion-exchange and gel-filtration fast protein liquid chromatography. The protein could also be isolated using rate zonal centrifugation in sucrose gradients followed by conventional ion-exchange on DEAE-cellulose. The active enzyme was composed of 44 and 41 kilodalton subunits. Antibodies to the activase polypeptides were produced in tumor-induced mouse ascites fluid and used as probes for activase on immunoblots of soluble proteins from a number of species. One or both of the activase polypeptides were recognized in all higher plant species examined including Arabidopsis thaliana, soybean, kidney bean, pea, tobacco, maize, oat, barley, celery, tomato, pigweed, purslane, dandelion, sorghum, and crabgrass. The polypeptides were not present in a mutant of Arabidopsis which is incapable of activating rubisco in vivo. The activase polypeptides were also detected in cell extracts of the green alga Chlamydomonas reinhardii. Activase activity, which had been demonstrated previously in wild-type Arabidopsis and in spinach, was measured in protoplast extracts of Nicotiana rustica. The results suggest that control of rubisco by activase may be an ubiquitous form of regulation in eucaryotic photosynthetic organisms.