Interaction between HLA-DM and HLA-DR involves regions that undergo conformational changes at lysosomal pH.
Antigenic peptide loading of major histocompatibility complex class II molecules is enhanced by lysosomal pH and catalyzed by the HLA-DM molecule. The physical mechanism behind the catalytic activity of DM was investigated by using time-resolved fluorescence anisotropy (TRFA) and fluorescence binding studies with the dye 8-anilino-1-naphthalenesulfonic acid (ANS). We demonstrate that the conformations of both HLA-DM and HLA-DR3, irrespective of the composition of bound peptide, are pH sensitive. Both complexes reversibly expose more nonpolar regions upon protonation. Interaction of DM with DR shields these hydrophobic domains from the aqueous environment, leading to stabilization of the DM and DR conformations. At lysosomal pH, the uncovering of additional hydrophobic patches leads to a more extensive DM-DR association. We propose that DM catalyzes class II peptide loading by stabilizing the low-pH conformation of DR, favoring peptide exchange. The DM-DR association involves a larger hydrophobic surface area with DR/class II-associated invariant chain peptides (CLIP) than with stable DR/peptide complexes, explaining the preferred association of DM with the former. The data support a release mechanism of DM from the DM-DR complex through reduction of the interactive surface, upon binding of class II molecules with antigenic peptide or upon neutralization of the DM-DR complex at the cell surface.
Ullrich
H J
HJ
Human Immunogenetics Laboratory, Imperial Cancer Research Fund, Holborn, London, United Kingdom.
Döring
K
K
Grüneberg
U
U
Jähnig
F
F
Trowsdale
J
J
van Ham
S M
SM
D000817:Anilino Naphthalenesulfonates; D002460:Cell Line; D005456:Fluorescent Dyes; D006681:HLA-D Antigens / Q000737:chemistry / Q000378:metabolism*; D006684:HLA-DR Antigens / Q000737:chemistry / Q000378:metabolism*; D000949:Histocompatibility Antigens Class II; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008247:Lysosomes / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; C416189:H2-M antigens; D006681:HLA-D Antigens; C084245:HLA-DM antigens; D006684:HLA-DR Antigens; D000949:Histocompatibility Antigens Class II; D011994:Recombinant Proteins; C027132:1-anilino-8-naphthalenesulfonate