title: The 70-kDa heat-shock protein/DnaK chaperone system is required for the productive folding of ribulose-biphosphate carboxylase subunits in Escherichia coli.
abstract: We have studied the in vivo requirements of the DnaK chaperone system for the folding of recombinant ribulose-bisphosphate carboxylase/oxygenase in Escherichia coli. Expression of functional dimeric or hexadecameric ribulose-bisphosphate carboxylase from different bacterial sources (including purple bacteria and cyanobacteria) was severely impaired in E. coli dnaK, dnaJ, or grpE mutants. These enzymes were synthesized mostly in soluble, fully enzymatically active forms in wild-type E. coli cells cultured in the temperature range 20-42 degrees C, but aggregated extensively in dnaK null mutants. Co-expression of dnaK, but not groESL, markedly reduced the aggregation of ribulose-bisphosphate carboxylase subunits in dnaK null mutants and restored the enzyme activity to levels found in isogenic wild-type strains. Ribulose-bisphosphate carboxylase expression in wild-type E. coli cells growing at 30 degrees C promoted an enhanced synthesis of stress proteins, apparently by sequestering DnaK from its negative regulatory role in this response. The overall results indicate that the DnaK chaperone system assists in vivo the folding pathway of ribulose-bisphosphate carboxylase large subunits, most probably at its very early stages.
authors.affiliation: Departamento de Microbiología, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Argentina.
authors.forename: S K
authors.initials: SK
authors.lastname: Checa
authors.forename: A M
authors.initials: AM
authors.lastname: Viale
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D018833:Chaperonins / Q000235:genetics / Q000378:metabolism; D000458:Cyanobacteria / Q000201:enzymology; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012273:Ribulose-Bisphosphate Carboxylase / Q000737:chemistry* / Q000378:metabolism; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C097970:GroESL protein, Bacteria; D018840:HSP70 Heat-Shock Proteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D018833:Chaperonins; C063842:dnaK protein, E coli; D012273:Ribulose-Bisphosphate Carboxylase