title: Chaperone-based procedure to increase yields of soluble recombinant proteins produced in E. coli.
abstract: BACKGROUND
  The overproduction of recombinant proteins in host cells often leads to their misfolding and aggregation. Previous attempts to increase the solubility of recombinant proteins by co-overproduction of individual chaperones were only partially successful. We now assessed the effects of combined overproduction of the functionally cooperating chaperone network of the E. coli cytosol on the solubility of recombinant proteins.
  
  
  RESULTS
  A two-step procedure was found to show the strongest enhancement of solubility. In a first step, the four chaperone systems GroEL/GroES, DnaK/DnaJ/GrpE, ClpB and the small HSPs IbpA/IbpB, were coordinately co-overproduced with recombinant proteins to optimize de novo folding. In a second step, protein biosynthesis was inhibited to permit chaperone mediated refolding of misfolded and aggregated proteins in vivo. This novel strategy increased the solubility of 70% of 64 different heterologous proteins tested up to 42-fold.
  
  
  CONCLUSIONS
  The engineered E. coli strains and the two-step procedure presented here led to a remarkable increase in the solubility of a various recombinant proteins and should be applicable to a wide range of target proteins produced in biotechnology.
authors.affiliation: EMBL Heidelberg, Heidelberg, Germany. ario.demarco@ifom-ieo-campus.it
authors.forename: Ario
authors.initials: A
authors.lastname: de Marco
authors.forename: Elke
authors.initials: E
authors.lastname: Deuerling
authors.forename: Axel
authors.initials: A
authors.lastname: Mogk
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Tomoyasu
authors.forename: Bernd
authors.initials: B
authors.lastname: Bukau
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D024861:Genetic Enhancement / Q000379:methods; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012995:Solubility
chemical_list: D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins