Molecular architecture of the TAP-associated MHC class I peptide-loading complex.
Tapasin organizes the peptide-loading complex (PLC) by recruiting peptide-receptive MHC class I (MHC-I) and accessory chaperones to the N-terminal regions of the TAP subunits TAP1 and TAP2. Despite numerous studies have shown that the formation of the PLC is essential to facilitate proper MHC-I loading, the molecular architecture of this complex is still highly controversial. We studied the stoichiometry of the PLC by blue native-PAGE in combination with Ab-shift assays and found that TAP/tapasin complexes exist at steady state as a mixture of two distinct oligomers of 350 and 450 kDa. Only the higher m.w. complex contains MHC-I and disulfide-linked tapasin/ER60 conjugates. Moreover, we show for the first time to our knowledge that the fully assembled PLC comprises two tapasin, two ER60, but only one complex of MHC-I and calreticulin. Based hereon we postulate that the TAP subunits alternate in the recruitment and loading of a single MHC-I.
Rufer
Elke
E
Friedrich-Loeffler-Institute, Federal Research Institute for Animal Health, Institute of Immunology, Tuebingen, Germany.
Leonhardt
Ralf M
RM
Knittler
Michael R
MR
D000071181:ATP Binding Cassette Transporter, Subfamily B, Member 2; D000071450:ATP Binding Cassette Transporter, Subfamily B, Member 3; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D017398:Alternative Splicing / Q000235:genetics; D001665:Binding Sites; D037282:Calreticulin / Q000378:metabolism; D002460:Cell Line; D019281:Dimerization; D004220:Disulfides / Q000378:metabolism; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D026901:Membrane Transport Proteins / Q000378:metabolism; D018832:Molecular Chaperones / Q000378:metabolism; D010455:Peptides / Q000378:metabolism*; D011485:Protein Binding; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
D000071181:ATP Binding Cassette Transporter, Subfamily B, Member 2; D000071450:ATP Binding Cassette Transporter, Subfamily B, Member 3; D018528:ATP-Binding Cassette Transporters; D037282:Calreticulin; D004220:Disulfides; D015395:Histocompatibility Antigens Class I; D026901:Membrane Transport Proteins; D018832:Molecular Chaperones; D010455:Peptides; C073119:TAP1 protein, human; C486172:Tap1 protein, rat; C489514:Tap2 protein, rat; C107762:tapasin; C071692:TAP2 protein, human; D010738:Type C Phospholipases; D019704:Protein Disulfide-Isomerases