The BSD 2 ortholog in Chlamydomonas reinhardtii is a polysome‐associated chaperone that co‐migrates on sucrose gradients with the rbcL transcript encoding the Rubisco large subunit

Summary The expression of the CO 2 ‐fixation enzyme ribulose‐bisphosphate carboxylase/oxygenase (Rubisco), which is affected by light, involves the cysteine‐rich protein bundle‐sheath defective‐2 ( BSD 2) that was originally identified in maize bundle‐sheath cells. We identified the BSD 2 ortholog in Chlamydomonas reinhardtii as a small protein (17 kDa) localized to the chloroplast. The algal BSD 2‐ortholog contains four CXXCXGXG DnaJ‐like elements, but lacks the other conserved domains of DnaJ. BSD 2 co‐migrated with the rbcL transcript on heavy polysomes, and both BSD 2 and rbcL m RNA shifted to the lighter fractions under oxidizing conditions that repress the translation of the Rubisco large subunit (RbcL). This profile of co‐migration supports the possibility that BSD2 is required for the de novo synthesis of RbcL. Furthermore, BSD 2 co‐migrated with the rbcL transcript in a C. reinhardtii premature‐termination mutant that encodes the first 60 amino acids of RbcL. In both strains, BSD 2 shared its migration profile with the rbcL transcript but not with psbA m RNA . The chaperone activity of BSD 2 was exemplified by its ability to prevent the aggregation of both citrate synthase ( CS ) and RbcL in vitro following their chemical denaturation. This activity did not depend on the presence of the thiol groups on BSD 2. In contrast, the activity of BSD 2 in preventing the precipitation of reduced β‐chains in vitro in the insulin turbidity assay was thiol‐dependent. We conclude that BSD 2 combines a chaperone ‘holdase’ function with the ability to interact with free thiols, with both activities being required to protect newly synthesized RbcL chains.