title: A specific, transmembrane interface regulates fibroblast activation protein (FAP) homodimerization, trafficking and exopeptidase activity.
abstract: Fibroblast activation protein (FAP) is a cell-surface serine protease which promotes invasiveness of certain epithelial cancers and is therefore a potential target for cancer drug development and delivery. Unlike dipeptidyl peptidase IV (DPPIV), FAP exhibits prolyl endopeptidase activity and is active as a homodimer with specificity for type I collagen. The mechanism that regulates FAP homodimerization and its relation to prolyl endopeptidase activity is not completely understood. Here, we investigate key residues in the FAP TM domain that may be significant for FAP homodimerization. Mutations to predicted TM interfacial residues (G10L, S14L, and A18L) comprising a small-X3-small motif reduced FAP TM-CYTO dimerization relative to wild type as measured using the AraTM assay, whereas predicted off-interface residues showed no significant change from wild type. The results implied that the predicted small-X3-small dimer interface affect stabilization of FAP TM-CYTO homodimerization. Compared with FAPwild-type, the interfacial TM residue G10L significantly decreased FAP endopeptidase activity more than 25%, and also reduced cell-surface versus intracellular expression relative to other interfacial residues S14L and A18L. Thus, our results suggest FAP dimerization is important for both trafficking and protease activity, and is dependent on a specific TM interface.
authors.affiliation: Program in Bioengineering, Lehigh University, Bethlehem, PA 18015, United States.
authors.forename: Benjamaporn
authors.initials: B
authors.lastname: Wonganu
authors.affiliation: Program in Bioengineering, Lehigh University, Bethlehem, PA 18015, United States; Department of Chemical and Biomolecular Engineering, Lehigh University, Bethlehem, PA 18015, United States. Electronic address: berger@lehigh.edu.
authors.forename: Bryan W
authors.initials: BW
authors.lastname: Berger
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D051777:AraC Transcription Factor / Q000235:genetics; D019281:Dimerization; D010450:Endopeptidases; D029968:Escherichia coli Proteins / Q000235:genetics; D018093:Gelatinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D000072417:Protein Domains; D021381:Protein Transport; D059748:Proteolysis; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000737:chemistry; D000099167:Fibroblast Activation Protein Alpha
chemical_list: D051777:AraC Transcription Factor; D010450:Endopeptidases; D029968:Escherichia coli Proteins; D018093:Gelatinases; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D012697:Serine Endopeptidases; D000099167:Fibroblast Activation Protein Alpha; C497371:AraC protein, E coli
keywords: Fibroblast activation protein; Homodimerization; Proteolytic activity; Transmembrane domain