Membrane topology and dimerization of the two subunits of the transporter associated with antigen processing reveal a three-domain structure

Presentation of peptides derived from cytosolic and nuclear proteins by MHC class I molecules requires their translocation across the membrane of the endoplasmic reticulum (ER) by a specialized ABC (ATP-binding cassette) tran sporter, TAP. To investigate the topology of the heterodimeric TAP complex, we constructed a set of C-terminal deletions for the TAP1 and TAP2 subunit s, We identified eight and seven transmembrane (TM) segments for TAP1 and T AP2, respectively. TAP1 has both its N and C terminus in the cytoplasm, whe reas TAP:! has its N terminus in the lumen of the ER. A putative TM pore co nsists of TM1-6 of TAP1 and, by analogy, TM1-5 of TAP2, Multiple PR-retenti on signals are present within this region, of which we positively identifie d TM1 of both TAP subunits. The N-terminal domain containing TM1-6 of TAP1 is sufficient for dimerization with TAP2, A second, independent dimerizatio n domain, located between the putative pore and the nucleotide-binding cass ette, lies within the cytoplasmic peptide-binding domains, which are anchor ed to the membrane via TM doublets 7/8 and 6/7 of TAP1 and TAP2, respective ly. We present a model in which TAP is composed of three subdomains: a TM p ore, a cytoplasmic peptide-binding pocket, and a nucleotide-binding domain.