title: Conformational state and receptor recognition of the C-terminal domain of human alpha(2)-macroglobulin after dissociation into half-molecules.
abstract: BACKGROUND
  Dissociation of native human alpha(2)-macroglobulin (alpha(2)M) by sodium thiocyanate generates stable half-molecules with intact thiol esters. Significant conformational changes occur by the dissociation, which are similar to those occurring by transformation from native to methylamine-treated alpha(2)-macroglobulin.
  
  
  METHODS
  The conformational state of the receptor-binding domain of the half-molecules was investigated by receptor binding and clearance studies, and by use of a panel of 11 monoclonal antibodies (mAbs) specific for the 18-kDa C-terminal receptor-binding fragment of alpha(2)-macroglobulin.
  
  
  RESULTS
  The half-molecules simultaneously express epitopes specific for native, as well as epitopes specific for transformed alpha(2)-macroglobulin. While it is possible to immunochemically discriminate between the different forms of tetrameric protein, the half-molecules retain a conformational state with no observed conformational changes in the C-terminal domain following cleavage of thiol esters or bait regions. The in vivo clearance rate in mice was consequently significantly slower for the half-molecules than for the tetrameric receptor-recognized forms of alpha(2)-macroglobulin. Furthermore, half-molecules demonstrate lower affinity for binding to mouse macrophages than methylamine-treated tetrameric alpha(2)-macroglobulin in competition studies.
  
  
  CONCLUSIONS
  It is suggested that contact zones are functionally important for mediating conformational switches, which result in trapping and exposure of the receptor-binding sites.
authors.affiliation: The Neuroimmunology Laboratory, The Center for Neuroscience, Copenhagen University Hospital, Section 9202, Rigshospitalet, Juliane Mariesvej 24, 2100 Copenhagen, Denmark. pejensen@pet.rh.dk
authors.forename: P E
authors.initials: PE
authors.lastname: Jensen
authors.forename: M
authors.initials: M
authors.lastname: Gunnarsson
authors.forename: T
authors.initials: T
authors.lastname: Stigbrand
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D006801:Humans; D066298:In Vitro Techniques; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008657:Metabolic Clearance Rate; D008744:Methylamines; D051379:Mice; D008807:Mice, Inbred BALB C; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011971:Receptors, Immunologic / Q000378:metabolism; D013861:Thiocyanates; D000511:alpha-Macroglobulins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008744:Methylamines; D011971:Receptors, Immunologic; D013861:Thiocyanates; D000511:alpha-Macroglobulins; C024553:sodium thiocyanate; C027451:methylamine