SPINACH CHLOROPLAST CPN21 CO-CHAPERONIN POSSESSES 2 FUNCTIONAL DOMAINS FUSED TOGETHER IN A TOROIDAL STRUCTURE AND EXHIBITS NUCLEOTIDE-DEPENDENT BINDING TO PLASTID CHAPERONIN-60

Chloroplasts contain a 21-kDa co-chaperonin polypeptide (cpn21) formed by two GroES-like domains fused together in tandem. Expression of a d ouble-domain spinach cpn21 in Escherichia coli groES mutant strains su pports growth of bacteriophages lambda and T5, and will also suppress a temperature-sensitive growth phenotype of a groES619 strain. Each do main of cpn21 expressed separately can function independently to suppo rt bacteriophage lambda growth, and the N-terminal domain will additio nally suppress the temperature-sensitive growth phenotype. These resul ts indicate that chloroplast cpn21 has two functional domains, either of which can interact with GroEL in vivo to facilitate bacteriophage m orphogenesis. Purified spinach cpn21 has a ring-like toroidal structur e and forms a stable complex with E. coli GroEL in the presence of ADP and is functionally interchangeable with bacterial GroES in the chape ronin-facilitated refolding of denatured ribulose-1,5-bisphosphate car boxylase. Cpn21 also inhibits the ATPase activity of GroEL. Cpn21 bind s with similar efficiency to both the alpha and beta subunits of spina ch cpn60 in the presence of adenine nucleotides, with ATP being more e ffective than ADP. The tandemly fused domains of cpn21 evolved early a nd are present in a wide range of photosynthetic eukaryotes examined, indicating a high degree of conservation of this structure in chloropl asts.