title: Probing the interface in a human co-chaperonin heptamer: residues disrupting oligomeric unfolded state identified.
abstract: BACKGROUND
  The co-chaperonin protein 10 (cpn10) assists cpn60 in the folding of nonnative polypeptides in a wide range of organisms. All known cpn10 molecules are heptamers of seven identical subunits that are linked together by beta-strand interactions at a large and flexible interface. Unfolding of human mitochondrial cpn10 in urea results in an unfolded heptameric state whereas GuHCl additions result in unfolded monomers. To address the role of specific interface residues in the assembly of cpn10 we prepared two point-mutated variants, in each case removing a hydrophobic residue positioned at the subunit-subunit interface.
  
  
  RESULTS
  Replacing valine-100 with a glycine (Val100Gly cpn10) results in a wild-type-like protein with seven-fold symmetry although the thermodynamic stability is decreased and the unfolding processes in urea and GuHCl both result in unfolded monomers. In sharp contrast, replacing phenylalanine-8 with a glycine (Phe8Gly cpn10) results in a protein that has lost the ability to assemble. Instead, this protein exists mostly as unfolded monomers.
  
  
  CONCLUSIONS
  We conclude that valine-100 is a residue important to adopt an oligomeric unfolded state but it does not affect the ability to assemble in the folded state. In contrast, phenylalanine-8 is required for both heptamer assembly and monomer folding and therefore this mutation results in unfolded monomers at physiological conditions. Despite the plasticity and large size of the cpn10 interface, our observations show that isolated interface residues can be crucial for both the retention of a heptameric unfolded structure and for subunit folding.
authors.affiliation: Chemistry Department, Tulane University, New Orleans, 70118 Louisiana, USA. guidry@tulane.edu
authors.forename: Jesse J
authors.initials: JJ
authors.lastname: Guidry
authors.forename: Frank
authors.initials: F
authors.lastname: Shewmaker
authors.forename: Karol
authors.initials: K
authors.lastname: Maskos
authors.forename: Samuel
authors.initials: S
authors.lastname: Landry
authors.forename: Pernilla
authors.initials: P
authors.lastname: Wittung-Stafshede
mesh_terms: D018835:Chaperonin 10 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019791:Guanidine / Q000494:pharmacology; D006801:Humans; D046911:Macromolecular Substances; D010649:Phenylalanine / Q000235:genetics; D017354:Point Mutation; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D013696:Temperature; D014508:Urea / Q000494:pharmacology; D014633:Valine / Q000235:genetics
chemical_list: D018835:Chaperonin 10; D046911:Macromolecular Substances; D021122:Protein Subunits; D010649:Phenylalanine; D014508:Urea; D014633:Valine; D019791:Guanidine