title: Functional asymmetry of the ATP-binding-cassettes of the ABC transporter TAP is determined by intrinsic properties of the nucleotide binding domains.
abstract: The ATP-binding-cassette (ABC) transporter associated with antigen processing (TAP) delivers peptides into the ER. TAP consists of two polypeptides (TAP1 and TAP2) each with an N-terminal transmembrane (TMD) and a C-terminal nucleotide binding domain (NBD). The two highly homologous NBDs of TAP show different nucleotide binding specificites, and identical mutations in the domains can have different effects on peptide transport. We asked whether this functional asymmetry of the NBDs is an intrinsic property or is imposed by the TMDs to which they are linked. To investigate the functional interdependence of the TAP domains, we created various TAP variants in which TMDs and/or NBDs were exchanged. All TAP variants except those with two TMDs of TAP1 could assemble. The TMDs did not affect the different nucleotide binding properties of the NBDs. The TAP variant with switched NBDs showed active peptide transport while the variants with pairs of identical NBDs or TMDs were inactive. Although both types of TMDs and NBDs have to be present for peptide transport they do not have to be assorted as in wild-type TAP. Thus, TAP domains seem to preserve functional autonomy despite their fusion into single polypeptide chains. We propose that the two NBDs act as nonequivalent 'modules' that directly determine the functional asymmetry of the included ATP-binding-cassettes. This provides a new insight into the function of NBDs and opens up new possibilities to investigate the molecular mechanism of the 'NBD engine' in ABC transporters.
authors.affiliation: Institute for Genetics, University of Cologne, Germany.
authors.forename: O
authors.initials: O
authors.lastname: Daumke
authors.forename: M R
authors.initials: MR
authors.lastname: Knittler
mesh_terms: 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*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D005434:Flow Cytometry; D006801:Humans; D026901:Membrane Transport Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014407:Tumor Cells, Cultured
chemical_list: D000071181:ATP Binding Cassette Transporter, Subfamily B, Member 2; D000071450:ATP Binding Cassette Transporter, Subfamily B, Member 3; D018528:ATP-Binding Cassette Transporters; D026901:Membrane Transport Proteins; D009711:Nucleotides; D011993:Recombinant Fusion Proteins; C073119:TAP1 protein, human; C486172:Tap1 protein, rat; C489514:Tap2 protein, rat; C071692:TAP2 protein, human; D000255:Adenosine Triphosphate; C043912:peptide permease