Empty and peptide-loaded class II major histocompatibility complex proteins produced by expression in Escherichia coli and folding in vitro

Protein Expr Purif. 1999 Feb;15(1):105-14. doi: 10.1006/prep.1998.0987.


The human class II major histocompatibility complex protein HLA-DR1 has been expressed in Escherichia coli as denatured alpha and beta subunits and folded in vitro to form the native structure. DR1 folding yields are 30-50% in the presence or absence of tight-binding antigenic peptides. The protein produced in this manner is soluble and monomeric with the expected apparent molecular weight. It reacts with conformation-sensitive anti-DR antibodies and exhibits peptide-dependent resistance to SDS-induced chain dissociation and to proteolysis as does the native protein. The observed peptide specificity and dissociation kinetics are similar to those of native DR produced in B-cells and finally the protein exhibits circular dichroism spectra and cooperative thermal denaturation as expected for a folded protein. We conclude that the recombinant DR1 has adopted the native fold. We have folded DR1 in the absence of peptide and isolated a soluble, peptide-free alphabeta-heterodimer. The empty DR1 can bind antigenic peptide but exhibits altered far UV-circular dichroism and thermal denaturation relative to the peptide-bound form.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Antibodies
  • Binding Sites
  • Cloning, Molecular / methods
  • Escherichia coli
  • HLA-DR1 Antigen / biosynthesis*
  • HLA-DR1 Antigen / chemistry*
  • HLA-DR1 Antigen / isolation & purification
  • Hot Temperature
  • Humans
  • Kinetics
  • Macromolecular Substances
  • Molecular Sequence Data
  • Peptide Fragments / chemistry*
  • Peptide Fragments / metabolism
  • Protein Conformation*
  • Protein Denaturation
  • Protein Folding*
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Thermodynamics


  • Antibodies
  • HLA-DR1 Antigen
  • Macromolecular Substances
  • Peptide Fragments
  • Recombinant Proteins