title: Expression and characterization of a variant of TACI (CRD2-shortTACIFc) in Pichia pastoris.
abstract: TACI is a member of the tumor necrosis factor receptor superfamily and serves as a key regulator of B cell function. The extracellular domain of a typical TNFR contains multiple copies of CRD, which bind in the monomermonomer interfaces of a trimeric ligand. TACI binds to two ligands, APRIL and BAFF, with high affinity and contains two CRD in its extracellular regions, while BCMA and BR3, contain a single or partial CRD for binding the two ligands. However, TACI can be classified as a single CRD receptor because the amino-terminal CRD1 doesn't contribute to ligand binding. To obtain a new variant of TACI possessing higher affinities for binding, we fused a repeat sequence of CRD2 to the N-terminus of the short form of TACI. The new APRIL antagonist peptide, CRD2-shortTACI-Fc, was designed based on the modeling 3-D complex structure of TACI and APRIL. As expected, the purified recombinant CRD2-shortTACI-Fc fusion protein could bind to APRIL in vitro and demonstrated dose-dependent inhibition of APRIL-induced proliferative activity in Raji cells. We found that CRD2-shortTACI-Fc, has a higher affinity for binding to ligands than short-TACI-Fc, which contains a single CRD2.
authors.affiliation: Department of Biochemistry and Molecular Biology, Key Laboratory of Molecular Medicine, the Ministry of Education, Shanghai Medical College, Shanghai, People’s Republiof China.
authors.forename: Rui
authors.initials: R
authors.lastname: Wang
authors.forename: Shiliang
authors.initials: S
authors.lastname: Zhou
authors.forename: Xiaomin
authors.initials: X
authors.lastname: Peng
authors.forename: Xin-Wen
authors.initials: XW
authors.lastname: Zhou
authors.forename: Zhi-Qun
authors.initials: ZQ
authors.lastname: Xie
authors.forename: Yuxiong
authors.initials: Y
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Mo
authors.forename: Min
authors.initials: M
authors.lastname: Yu
mesh_terms: D000595:Amino Acid Sequence; D019677:Artificial Gene Fusion; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D015398:Signal Transduction; D053303:Transmembrane Activator and CAML Interactor Protein / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D053300:Tumor Necrosis Factor Ligand Superfamily Member 13 / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
chemical_list: D011993:Recombinant Fusion Proteins; C108467:TNFRSF13B protein, human; D053303:Transmembrane Activator and CAML Interactor Protein; D053300:Tumor Necrosis Factor Ligand Superfamily Member 13