Hydrophobic surfaces that are hidden in chaperonin Cpn60 can be exposed by formation of assembly-competent monomers or by ionic perturbation of the oligomer

J Biol Chem. 1995 Jan 27;270(4):1535-42.


The oligomeric form (14-mer) of the chaperonin protein, Cpn60 (GroEL) from Eschericia coli, displays restricted hydrophobic surfaces and binds tightly one to two molecules of the fluorescent hydrophobic reporter, 1,1'-bi(4-anilino)naphthalene-5,5'-disulfonic acid (bisANS). The 14-mer is resistant to proteolysis by chymotrypsin, and none of the three sulfhydryl groups/monomer react with 6-iodoacetamidofluorescein. When monomers of Cpn60 that are folded and competent to participate in protein folding are formed by low concentrations of urea (< 2.5 M), the hydrophobic exposure increases to accommodate approximately 14 molecules of bisANS/14-mer, the binding affinity for bisANS decreases, and 1 sulfhydryl group/monomer reacts with 6-iodoacetamidofluorescein. These monomers display limited proteolysis by chymotrypsin at several points within a hydrophobic sequence centered around residue 250 to produce a relatively stable N-terminal fragment (approximately = to 26 kDa) and a partially overlapping C-terminal fragment (approximately = to 44 kDa). The exposure of hydrophobic surfaces is facilitated by ATPMg. Ions increase hydrophobic exposure more effectively than urea without dissociation of Cpn60. For example, subdenaturing concentrations of guanidinium chloride (< or = 0.75 M) or the stabilizing salt, guanidinium sulfate, as well as NaCl or KCl are effective. The trivalent cation, spermidine, induces maximum exposure at 5 mM. The results suggest that hydrophobic surfaces can be involved in stabilizing the oligomer and/or in binding proteins to be folded, and they are consistent with suggestions that amphiphilic structures, presenting hydrophobic surfaces within a charged context, would be particularly effective in binding to Cpn60.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Anilino Naphthalenesulfonates
  • Chaperonin 60 / chemistry*
  • Chaperonin 60 / metabolism
  • Chymotrypsin
  • Escherichia coli
  • Fluorescent Dyes
  • Guanidine
  • Guanidines / pharmacology
  • Macromolecular Substances
  • Osmolar Concentration
  • Peptide Fragments / chemistry
  • Peptide Fragments / isolation & purification
  • Protein Binding
  • Protein Conformation*
  • Protein Denaturation
  • Protein Folding
  • Spectrometry, Fluorescence
  • Surface Properties
  • Ultracentrifugation
  • Urea / pharmacology


  • Anilino Naphthalenesulfonates
  • Chaperonin 60
  • Fluorescent Dyes
  • Guanidines
  • Macromolecular Substances
  • Peptide Fragments
  • 5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate)
  • Urea
  • Chymotrypsin
  • Guanidine