Expression of the Arabidopsis thaliana AtJ2 cochaperone protein in Pichia pastoris

A vector was constructed for intracellular expression of the Arabidopsis th aliana DnaJ homologue AtJ2 in the methylotrophic yeast Pichia pastoris. The vector includes DNA encoding an amino-terminal histidine-tag, to simplify protein purification. Shake-flask cultures could be induced to produce appr oximately 250 mg/L of AtJ2. Purified recombinant AtJ2 was able to stimulate the ATPase activities of both the Escherichia coli and Zea mays cytoplasmi c Stress70 chaperone proteins five- to ninefold. The carboxy terminus of At J2 is -CAQQ, a protein farnesylation motif. When transformed P. pastoris wa s induced to synthesize AtJ2 in the presence of [H-3]mevalonolactone, radio activity was incorporated into the protein, suggesting farnesylation, (C) 2 000 Academic Press.